EUROIMMUN ANTI-NRNP/SM ELISA (IGG)

• K Number: K123261
• Device Name: EUROIMMUN ANTI-NRNP/SM ELISA (IGG)
• Manufacturer: EUROIMMUN US
• Date Cleared: 2013-06-12 ( 237 days)
• Product Code: LKO LJM LKP LLL MQA
• Regulation Number: 866.5100

Intended Use

The EUROIMMUN Anti-nRNP/Sm ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against nRNP/Sm in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of mixed connective tissue diseases and systemic lupus erythematosus, in conjunction with other laboratory and clinical findings. The EUROIMMUN Anti-Sm ELISA (IgG) test kit is intended for the qualitative of IgG class autoantibodies against Sm in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of systemic lupus erythematosus, in conjunction with other laboratory and clinical findings. The EUROIMMUN Anti-SS-A ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against SS-A in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of Sjögren's syndrome and systemic lupus erythematosus, in conjunction with other laboratory and clinical findings. The EUROIMMUN Anti-SS-B ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against SS-B in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of Sjögren's syndrome and systemic lupus erythematosus, in conjunction with other laboratory and clinical findings. The EUROIMMUN Anti-Scl-70 ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against Scl-70 in human serum and plasma (EDTA, Li-heparin, Citrate), It is used as an aid in the diagnosis of progressive systemic sclerosis, in conjunction with other laboratory and clinical findings. The EUROIMMUN Anti-Centromeres ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against Centromeres in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of limited form of progressive systemic sclerosis (CREST syndrome), in conjunction with other laboratory and clinical findings. The EUROIMMUN Anti-Jo-1 ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against Jo-1 in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of polymyositis and dermatomyositis, in conjunction with other laboratory and clinical findings. The EUROIMMUN Anti-ribosomal P-proteins ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against ribosomal P-proteins in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of systemic lupus erythematosus, in conjunction with other laboratory and clinical findings.

Device Description

The EUROIMMUN Anti-nRNP/Sm ELISA (IgG) consists of a microwell ELISA plate coated with nRNP/Sm antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution. The EUROIMMUN Anti-Sm ELISA (IgG) consists of a microwell ELISA plate coated with Sm antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution. The EUROIMMUN Anti-SS-A ELISA (IgG) consists of a microwell ELISA plate coated with SS-A antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate. TMB chromogen/substrate solution and stop solution. The EUROIMMUN Anti-SS-B ELISA (IgG) consists of a microwell ELISA plate coated with SS-B antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution. The EUROIMMUN Anti-Scl-70 ELISA (IgG) consists of a microwell ELISA plate coated with Scl-70 antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution. The EUROIMMUN Anti-Centromeres ELISA (IgG) consists of a microwell ELISA plate coated with Centromeres antigen, calibrator, positive control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution. The EUROIMMUN Anti-Jo-1 ELISA (IgG) consists of a microwell ELISA plate coated with Jo-1 antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution. The EUROIMMUN Anti-ribosomal P-proteins ELISA (IgG) consists of a microwell ELISA plate coated with ribosomal P-proteins antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution.

Predicate Devices

K922833, K922831, K922830, K922832, K924898, KOO3AEa, K922832, K981237

Reference Devices

K922833, K922831, K922830, K922832, K924898, KOO3AEa, K922832, K981237

AI/ML (Artificial Intelligence / Machine Learning)

No
The device description and intended use clearly describe a traditional ELISA assay kit for detecting autoantibodies, with no mention of AI or ML components.

Therapeutic

No.

This device is an in vitro diagnostic (IVD) test kit used to aid in the diagnosis of various autoimmune diseases by detecting autoantibodies. It does not directly treat or alleviate a condition.

Diagnostic

Yes

The "Intended Use / Indications for Use" section explicitly states that the test kits are "used as an aid in the diagnosis" of various diseases, directly indicating their diagnostic purpose.

SaMD (Software as a Medical Device)

No

The device description clearly outlines physical components of an ELISA test kit, including microwell plates, reagents, and solutions, indicating it is a hardware-based diagnostic test, not software only.

IVD (In Vitro Diagnostic)

Yes, this device is an IVD (In Vitro Diagnostic).

Here's why:

• Intended Use: The intended use clearly states that the test kits are for the "qualitative determination of IgG class autoantibodies" in "human serum and plasma". This indicates that the device is used to examine specimens derived from the human body.
• Purpose: The intended use also states that the tests are used "as an aid in the diagnosis" of various diseases (mixed connective tissue diseases, systemic lupus erythematosus, Sjögren's syndrome, progressive systemic sclerosis, polymyositis, and dermatomyositis). This aligns with the definition of an IVD, which is used to provide information for the diagnosis, prevention, monitoring, treatment, or alleviation of disease.
• Device Description: The device description details the components of the test kits, which are designed to perform an ELISA (Enzyme-Linked Immunosorbent Assay) on biological samples. This is a common method used in IVD testing.
• Performance Studies: The document includes detailed performance studies using clinically characterized samples, demonstrating that the device is being evaluated for its ability to detect specific analytes in human specimens for diagnostic purposes.
• Predicate Devices: The mention of predicate devices with K numbers (which are FDA premarket notification numbers for medical devices, including IVDs) further confirms that these types of tests are regulated as IVDs.

All these factors indicate that the EUROIMMUN ELISA test kits described are intended for in vitro diagnostic use.

PCCP (Predetermined Change Control Plan) Authorized

N/A

Overview

Intended Use / Indications for Use

The EUROIMMUN Anti-nRNP/Sm ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against nRNP/Sm in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of mixed connective tissue diseases and systemic lupus erythematosus, in conjunction with other laboratory and clinical findings.

The EUROIMMUN Anti-Sm ELISA (IgG) test kit is intended for the qualitative of IgG class autoantibodies against Sm in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of systemic lupus erythematosus, in conjunction with other laboratory and clinical findings.

The EUROIMMUN Anti-SS-A ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against SS-A in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of Sjögren's syndrome and systemic lupus erythematosus, in conjunction with other laboratory and clinical findings.

The EUROIMMUN Anti-SS-B ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against SS-B in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of Sjögren's syndrome and systemic lupus erythematosus, in conjunction with other laboratory and clinical findings.

The EUROIMMUN Anti-Scl-70 ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against Scl-70 in human serum and plasma (EDTA, Li-heparin, Citrate), It is used as an aid in the diagnosis of progressive systemic sclerosis, in conjunction with other laboratory and clinical findings.

The EUROIMMUN Anti-Centromeres ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against Centromeres in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of limited form of progressive systemic sclerosis (CREST syndrome), in conjunction with other laboratory and clinical findings.

The EUROIMMUN Anti-Jo-1 ELISA (IqG) test kit is intended for the qualitative determination of IgG class autoantibodies against Jo-1 in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of polymyositis and dermatomyositis, in conjunction with other laboratory and clinical findings.

The EUROIMMUN Anti-ribosomal P-proteins ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against ribosomal P-proteins in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of systemic lupus erythematosus, in conjunction with other laboratory and clinical findings.

Product codes

LKO, LKP, LLL, LJM, MQA

Device Description

The EUROIMMUN Anti-nRNP/Sm ELISA (IgG) consists of a microwell ELISA plate coated with nRNP/Sm antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution.

The EUROIMMUN Anti-Sm ELISA (IgG) consists of a microwell ELISA plate coated with Sm antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution.

The EUROIMMUN Anti-SS-A ELISA (IgG) consists of a microwell ELISA plate coated with SS-A antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate. TMB chromogen/substrate solution and stop solution.

The EUROIMMUN Anti-SS-B ELISA (IgG) consists of a microwell ELISA plate coated with SS-B antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution.

The EUROIMMUN Anti-Scl-70 ELISA (IgG) consists of a microwell ELISA plate coated with Scl-70 antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution.

The EUROIMMUN Anti-Centromeres ELISA (IgG) consists of a microwell ELISA plate coated with Centromeres antigen, calibrator, positive control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution.

The EUROIMMUN Anti-Jo-1 ELISA (IgG) consists of a microwell ELISA plate coated with Jo-1 antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution.

The EUROIMMUN Anti-ribosomal P-proteins ELISA (IgG) consists of a microwell ELISA plate coated with ribosomal P-proteins antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution.

Mentions image processing

Not Found

Mentions AI, DNN, or ML

Not Found

Input Imaging Modality

Not Found

Anatomical Site

Not Found

Indicated Patient Age Range

Not Found

Intended User / Care Setting

For prescription use only.

Description of the training set, sample size, data source, and annotation protocol

Not Found

Description of the test set, sample size, data source, and annotation protocol

Not Found

Summary of Performance Studies (study type, sample size, AUC, MRMC, standalone performance, key results)

Anti-nRNP/Sm ELISA (IgG)
Precision/Reproducibility:
Intra-assay reproducibility based on 20 determinations and inter-assay reproducibility on 40 determinations performed in 10 different runs on 5 days with 2 runs per day.
Expected result was positive for samples 1-4 and negative for samples 5-6.
Intra-assay % positive/negative matched expected for all samples.
Inter-assay % positive/negative matched expected for all samples.
Lot to lot reproducibility: 6* and 11** samples (n lots x 1 run).
Expected result was positive for samples 1-3, 6-7, 8-11 and negative for samples 4, 5.
Lot to lot reproducibility % positive/negative matched expected for all samples.

Comparison with predicate device:
Study performed using 287 clinically characterized samples (52 MCTD, 69 SLE, 51 Sjögren's syndrome, 15 systemic sclerosis, 15 fibromyalgia, 35 RA, 30 borreliosis and 20 healthy).
Negative agreement: 196/196 = 100.0% (95% C.I.: 98.1% - 100.0%).
Positive agreement: 83/91 = 91.2% (95% C.I.: 83.4% - 96.1%).
Overall agreement: 279/287 = 97.2% (95% C.I.: 94.6% - 98.8%).

Matrix comparison:
Usability of plasma investigated using sample pairs of serum and corresponding plasma (EDTA, Li-heparin, Citrate).
Regression equation for serum to plasma was near ideal correlation (intercept 0; slope 1.0).
Coefficient of determination R²: EDTA plasma 0.9988, Li-heparin plasma 0.9929, Citrate plasma 0.9915.
Mean %recovery: EDTA plasma 101%, Li-heparin plasma 103%, Citrate plasma 105%.

Clinical studies:
Total 1046 clinically characterized samples (65 from MCTD patients, 404 from SLE patients, 151 from myositis patients and 426 from control groups) investigated.
Clinical sensitivity in MCTD: 100.0% (95% C.I.: 94.5 - 100.0%).
Clinical sensitivity in SLE: 23.3% (95% C.I.: 19.2 - 27.7%).
Clinical specificity: 99.3% (95% C.I.: 98.0 - 99.9%).

Expected values/Reference range:
Panel of 200 samples from apparently healthy blood donors (120 men and 80 women with an average age of 40 y; age range: 19 - 68 y).
Prevalence: 0.5%.
Lowest value: 0.1, Highest value: 1.3, Mean value: 0.1, Std deviation: 0.09.

Anti-Sm ELISA (IgG)
Precision/Reproducibility:
Intra-assay reproducibility based on 20 determinations and inter-assay reproducibility on 40 determinations performed in 10 different runs on 5 days with 2 runs per day.
Intra-assay results were consistent with expected positive/negative classifications.
Inter-assay results showed excellent reproducibility with some minor variations for sample 5 (2.5% positive, 97.5% negative) but generally consistent with expected.
Lot to lot reproducibility was investigated using different lots with QC samples. Results showed good consistency with expected outcomes across different samples and lots.

Comparison studies:
Method comparison with predicate device (Inova Quanta Lite Sm ELISA) using 294 clinically characterized samples.
Negative agreement: 245/250 = 98.0% (95% C.I.: 95.4% - 99.3%).
Positive agreement: 37/44 = 84.1% (95% C.I.: 69.9% - 93.4%).
Overall agreement: 282/294 = 95.9% (95% C.I.: 93.0% - 97.9%).

Matrix comparison:
Usability of plasma (EDTA, Li-heparin, Citrate) compared to serum.
Regression equations were near ideal correlation; coefficients of determination > 0.99.
Mean %recovery: EDTA plasma 102%, Li-heparin plasma 99%, Citrate plasma 103%.
Range of %recovery: EDTA plasma 94-115%, Li-heparin plasma 90-111%, Citrate plasma 89-118%.

Clinical studies:
1036 clinically characterized samples (414 from SLE patients and 622 from control groups).
Clinical sensitivity for SLE: 11.4% (95% C.I.: 8.5 - 14.8%).
Clinical specificity for control groups: 99.0% (95% C.I.: 97.9 - 99.6%).

Expected values/Reference range:
Panel of 200 healthy blood donors (120 men, 80 women; average age 40 y, range 19-68 y).
Prevalence: 0.0% (0 positives, 200 negatives).
Ratio: Lowest 0.1, Highest 0.3, Mean 0.1, Std deviation 0.02.

Anti-SS-A ELISA (IgG)
Precision/Reproducibility:
Intra-assay reproducibility (n=20) and Inter-assay reproducibility (n=40) results for 6 samples. All samples showed 100% positive or negative detection matching expected results.
Lot to lot reproducibility showed consistent expected results across different lots and samples.

Comparison studies:
Method comparison with predicate device (Inova Quanta Lite SS-A ELISA) using 305 clinically characterized samples.
Negative agreement: 177 / 180 = 98.3% (95% C.I.: 95.2% - 99.7%).
Positive agreement: 116 / 125 = 92.8% (95% C.I.: 86.8% - 96.7%).
Overall agreement: 293 / 305 = 96.1% (95% C.I.: 93.2% - 98.0%).

Matrix comparison:
Usability of plasma (EDTA, Li-heparin, Citrate) compared to serum (n=16 each).
Regression equations were near ideal correlation; coefficients of determination R² > 0.99.
Mean %recovery: EDTA plasma 97%, Li-heparin plasma 95%, Citrate plasma 98%.
Range of %recovery: EDTA plasma 90-102%, Li-heparin plasma 85-102%, Citrate plasma 93-104%.

Clinical studies:
1026 clinically characterized samples (88 Sjögren's syndrome, 404 SLE, 534 control groups).
Clinical sensitivity for Sjögren's syndrome: 73.9% (95% C.I.: 63.4 - 82.7%).
Clinical sensitivity for SLE: 40.6% (95% C.I.: 35.8 - 45.6%).
Clinical specificity for control groups: 94.8% (95% C.I.: 92.5 - 96.5%).

Expected values/Reference range:
Panel of 200 healthy blood donors (120 men, 80 women; average age 40 y, range 19-68 y).
Prevalence: 1.0% (2 positives, 198 negatives).
Ratio: Lowest 0.0, Highest 4.4, Mean 0.1, Std deviation 0.33.

Anti-SS-B ELISA (IgG)
Precision/Reproducibility:
Intra-assay reproducibility (n=20) and Inter-assay reproducibility (n=40) results across 6 samples showed 100% positive or negative matching expected results.
Lot to lot reproducibility showed consistent expected results across different lots and samples.

Comparison studies:
Method comparison with predicate device (Inova Quanta Lite SS-B ELISA) using 275 clinically characterized samples.
Negative agreement: 189 / 190 = 99.5% (95% C.I.: 97.1% - 100.0%).
Positive agreement: 82 / 85 = 96.5% (95% C.I.: 90.0% - 99.3%).
Overall agreement: 271 / 275 = 98.5% (95% C.I.: 96.3% - 99.6%).

Matrix comparison:
Usability of plasma (EDTA, Li-heparin, Citrate) compared to serum using n=16 samples.
Regression equations were near ideal correlation; coefficients of determination R² > 0.99.
Mean %recovery: EDTA plasma 100%, Li-heparin plasma 105%, Citrate plasma 101%.
Range of %recovery: EDTA plasma 78-116%, Li-heparin plasma 98-116%, Citrate plasma 81-110%.

Clinical studies:
1026 clinically characterized samples (88 Sjögren's syndrome, 404 SLE, 534 control groups).
Clinical sensitivity for Sjögren's syndrome: 39.8% (95% C.I.: 29.5 - 50.8%).
Clinical sensitivity for SLE: 13.9% (95% C.I.: 10.6 - 17.6%).
Clinical specificity for control groups: 98.1% (95% C.I.: 96.6 - 99.1%).

Expected values/Reference range:
Panel of 200 healthy blood donors (120 men, 80 women; average age 40 y, range 19-68 y).
Prevalence: 0.0% (0 positives, 200 negatives).
Ratio: Lowest 0.0, Highest 0.2, Mean 0.0, Std deviation 0.02.

Anti-Scl-70 ELISA (IgG)
Precision/Reproducibility:
Intra-assay reproducibility (n=20) and Inter-assay reproducibility (n=40) results for 6 samples. All samples showed 100% positive or negative detection matching expected results.
Lot to lot reproducibility showed consistent expected results across different lots and samples.

Comparison studies:
Method comparison with predicate device (Inova Quanta Lite Scl-70 ELISA) using 309 clinically characterized samples.
Negative agreement: 184 / 184 = 100.0% (95% C.I.: 98.0% - 100.0%).
Positive agreement: 125 / 125 = 100.0% (95% C.I.: 97.1% - 100.0%).
Overall agreement: 309 / 309 = 100.0% (95% C.I.: 98.8% - 100.0%).

Matrix comparison:
Usability of plasma (EDTA, Li-heparin, Citrate) compared to serum using n=16 samples.
Regression equations were near ideal correlation; coefficients of determination R² > 0.98.
Mean %recovery: EDTA plasma 98%, Li-heparin plasma 101%, Citrate plasma 96%.
Range of %recovery: EDTA plasma 86-107%, Li-heparin plasma 92-113%, Citrate plasma 87-103%.

Clinical studies:
909 clinically characterized samples (280 systemic sclerosis, 629 control groups).
Clinical sensitivity for systemic sclerosis: 23.2% (95% C.I.: 18.4 - 28.6%).
Clinical sensitivity for diffuse systemic sclerosis: 59.4% (95% C.I.: 48.9 - 69.3%).
Clinical sensitivity for limited systemic sclerosis: 5.3% (95% C.I.: 2.0 - 11.2%).
Clinical specificity for control groups: 99.8% (95% C.I.: 99.1 - 100.0%).

Expected values/Reference range:
Panel of 200 healthy blood donors (120 men, 80 women; average age 40 y, range 19-68 y).
Prevalence: 0.0% (0 positives, 200 negatives).
Ratio: Lowest 0.0, Highest 0.1, Mean 0.0, Std deviation 0.01.

Anti-Centromeres ELISA (IgG)
Precision/Reproducibility:
Intra-assay reproducibility (n=20) and Inter-assay reproducibility (n=40) results for 6 samples. All samples showed 100% positive or negative detection matching expected results.
Lot to lot reproducibility showed consistent expected results across different lots and samples.

Comparison studies:
Method comparison with predicate device (Inova Quanta Lite Centromere ELISA) using 297 clinically characterized samples.
Negative agreement: 219 / 219 = 100.0% (95% C.I.: 98.3% - 100.0%).
Positive agreement: 73 / 78 = 93.6% (95% C.I.: 85.7% - 97.9%).
Overall agreement: 292 / 297 = 98.3% (95% C.I.: 96.1% - 99.5%).

Matrix comparison:
Usability of plasma (EDTA, Li-heparin, Citrate) compared to serum using n=16 samples.
Regression equations were near ideal correlation; coefficients of determination R² > 0.99.
Mean %recovery: EDTA plasma 98%, Li-heparin plasma 93%, Citrate plasma 96%.
Range of %recovery: EDTA plasma 84-108%, Li-heparin plasma 83-101%, Citrate plasma 88-107%.

Clinical studies:
877 clinically characterized samples (280 systemic sclerosis, 597 control groups).
Clinical sensitivity for systemic sclerosis: 37.5% (95% C.I.: 31.8 - 43.5%).
Clinical sensitivity for diffuse systemic sclerosis: 7.3% (95% C.I.: 3.0 - 14.4%).
Clinical sensitivity for limited systemic sclerosis: 74.3% (95% C.I.: 65.3 - 82.1%).
Clinical specificity for control groups: 99.0% (95% C.I.: 97.8 - 99.6%).

Expected values/Reference range:
Panel of 200 healthy blood donors (120 men, 80 women; average age 40 y, range 19-68 y).
Prevalence: 0.5% (1 positive, 199 negatives).
Ratio: Lowest 0.0, Highest 3.0, Mean 0.1, Std deviation 0.21.

Anti-Jo-1 ELISA (IgG)
Precision/Reproducibility:
Intra-assay reproducibility (n=20) and Inter-assay reproducibility (n=40) results for 6 samples. All samples showed 100% positive or negative detection matching expected results.
Lot to lot reproducibility showed consistent expected results across different lots and samples.

Comparison studies:
Method comparison with predicate device (Inova Quanta Lite Jo-1 ELISA) using 297 clinically characterized samples.
Negative agreement: 231 / 232 = 99.6% (95% C.I.: 97.6% - 100.0%).
Positive agreement: 64 / 65 = 98.5% (95% C.I.: 91.7% - 100.0%).
Overall agreement: 295 / 297 = 99.3% (95% C.I.: 97.6% - 99.9%).

Matrix comparison:
Usability of plasma (EDTA, Li-heparin, Citrate) compared to serum using n=15 samples.
Regression equations were near ideal correlation; coefficients of determination R² > 0.98.
Mean %recovery: EDTA plasma 101%, Li-heparin plasma 95%, Citrate plasma 103%.
Range of %recovery: EDTA plasma 85-117%, Li-heparin plasma 87-113%, Citrate plasma 94-114%.

Clinical studies:
876 clinically characterized samples (177 myositis, 699 control groups).
Clinical sensitivity for myositis: 18.6% (95% C.I.: 13.2 - 25.2%).
Clinical specificity for control groups: 99.6% (95% C.I.: 98.8 - 99.9%).

Expected values/Reference range:
Panel of 200 healthy blood donors (120 men, 80 women; average age 40 y, range 19-68 y).
Prevalence: 0.0% (0 positives, 200 negatives).
Ratio: Lowest 0.0, Highest 0.2, Mean 0.1, Std deviation 0.03.

Anti-ribosomal P-proteins ELISA (IgG)
Precision/Reproducibility:
Intra-assay reproducibility (n=20) and Inter-assay reproducibility (n=40) results for 6 samples. Majority of samples showed 100% positive/negative matching expected results, with one sample in inter-assay showing 2.5% positive/97.5% negative for an expected negative sample.
Lot to lot reproducibility showed consistent expected results across different lots and samples.

Comparison studies:
Method comparison with predicate device (Inova Quanta Lite Ribosomal P ELISA) using 243 clinically characterized samples.
Negative agreement: 205 / 215 = 95.3% (95% C.I.: 91.6% - 97.7%).
Positive agreement: 28 / 28 = 100.0% (95% C.I.: 87.7% - 100.0%).
Overall agreement: 233 / 243 = 95.9% (95% C.I.: 92.6% - 98.0%).

Matrix comparison:
Usability of plasma (EDTA, Li-heparin, Citrate) compared to serum using n=16 samples.
Regression equations were near ideal correlation; coefficients of determination R² > 0.99.
Mean %recovery: EDTA plasma 97%, Li-heparin plasma 98%, Citrate plasma 95%.
Range of %recovery: EDTA plasma 79-107%, Li-heparin plasma 80-107%, Citrate plasma 85-107%.

Clinical studies:
876 clinically characterized samples (376 SLE, 500 control groups).
Clinical sensitivity for SLE: 5.3% (95% C.I.: 3.3 - 8.1%).
Clinical specificity for control groups: 99.2% (95% C.I.: 98.0 - 99.8%).

Expected values/Reference range:
Panel of 150 healthy blood donors (79 men, 71 women; average age 38 y, range 18-67 y).
Prevalence: 0.0% (0 positives, 150 negatives).
Ratio: Lowest 0.0, Highest 0.7, Mean 0.1, Std deviation 0.08.

Key Metrics (Sensitivity, Specificity, PPV, NPV, etc.)

Anti-nRNP/Sm ELISA (IgG)
Clinical sensitivity in MCTD: 100.0% (95% C.I.: 94.5 - 100.0%)
Clinical sensitivity in SLE: 23.3% (95% C.I.: 19.2 - 27.7%)
Clinical specificity: 99.3% (95% C.I.: 98.0 - 99.9%)
Negative agreement: 100.0% (95% C.I.: 98.1% - 100.0%)
Positive agreement: 91.2% (95% C.I.: 83.4% - 96.1%)
Overall agreement: 97.2% (95% C.I.: 94.6% - 98.8%)

Anti-Sm ELISA (IgG)
Negative agreement: 98.0% (95% C.I.: 95.4% - 99.3%)
Positive agreement: 84.1% (95% C.I.: 69.9% - 93.4%)
Overall agreement: 95.9% (95% C.I.: 93.0% - 97.9%)
Clinical sensitivity for SLE: 11.4% (95% C.I.: 8.5 - 14.8%)
Clinical specificity for control groups: 99.0% (95% C.I.: 97.9 - 99.6%)

Anti-SS-A ELISA (IgG)
Negative agreement: 98.3% (95% C.I.: 95.2% - 99.7%)
Positive agreement: 92.8% (95% C.I.: 86.8% - 96.7%)
Overall agreement: 96.1% (95% C.I.: 93.2% - 98.0%)
Clinical sensitivity for Sjögren's syndrome: 73.9% (95% C.I.: 63.4 - 82.7%)
Clinical sensitivity for SLE: 40.6% (95% C.I.: 35.8 - 45.6%)
Clinical specificity for control groups: 94.8% (95% C.I.: 92.5 - 96.5%)

Anti-SS-B ELISA (IgG)
Negative agreement: 99.5% (95% C.I.: 97.1% - 100.0%)
Positive agreement: 96.5% (95% C.I.: 90.0% - 99.3%)
Overall agreement: 98.5% (95% C.I.: 96.3% - 99.6%)
Clinical sensitivity for Sjögren's syndrome: 39.8% (95% C.I.: 29.5 - 50.8%)
Clinical sensitivity for SLE: 13.9% (95% C.I.: 10.6 - 17.6%)
Clinical specificity for control groups: 98.1% (95% C.I.: 96.6 - 99.1%)

Anti-Scl-70 ELISA (IgG)
Negative agreement: 100.0% (95% C.I.: 98.0% - 100.0%)
Positive agreement: 100.0% (95% C.I.: 97.1% - 100.0%)
Overall agreement: 100.0% (95% C.I.: 98.8% - 100.0%)
Clinical sensitivity for systemic sclerosis: 23.2% (95% C.I.: 18.4 - 28.6%)
Clinical sensitivity for diffuse systemic sclerosis: 59.4% (95% C.I.: 48.9 - 69.3%)
Clinical sensitivity for limited systemic sclerosis: 5.3% (95% C.I.: 2.0 - 11.2%)
Clinical specificity for control groups: 99.8% (95% C.I.: 99.1 - 100.0%)

Anti-Centromeres ELISA (IgG)
Negative agreement: 100.0% (95% C.I.: 98.3% - 100.0%)
Positive agreement: 93.6% (95% C.I.: 85.7% - 97.9%)
Overall agreement: 98.3% (95% C.I.: 96.1% - 99.5%)
Clinical sensitivity for systemic sclerosis: 37.5% (95% C.I.: 31.8 - 43.5%)
Clinical sensitivity for diffuse systemic sclerosis: 7.3% (95% C.I.: 3.0 - 14.4%)
Clinical sensitivity for limited systemic sclerosis: 74.3% (95% C.I.: 65.3 - 82.1%)
Clinical specificity for control groups: 99.0% (95% C.I.: 97.8 - 99.6%)

Anti-Jo-1 ELISA (IgG)
Negative agreement: 99.6% (95% C.I.: 97.6% - 100.0%)
Positive agreement: 98.5% (95% C.I.: 91.7% - 100.0%)
Overall agreement: 99.3% (95% C.I.: 97.6% - 99.9%)
Clinical sensitivity for myositis: 18.6% (95% C.I.: 13.2 - 25.2%)
Clinical specificity for control groups: 99.6% (95% C.I.: 98.8 - 99.9%)

Anti-ribosomal P-proteins ELISA (IgG)
Negative agreement: 95.3% (95% C.I.: 91.6% - 97.7%)
Positive agreement: 100.0% (95% C.I.: 87.7% - 100.0%)
Overall agreement: 95.9% (95% C.I.: 92.6% - 98.0%)
Clinical sensitivity for SLE: 5.3% (95% C.I.: 3.3 - 8.1%)
Clinical specificity for control groups: 99.2% (95% C.I.: 98.0 - 99.8%)

Predicate Device(s)

Inova Quanta Lite RNP ELISA K922833, Inova Quanta Lite Sm ELISA K922831, Inova Quanta Lite SS-A ELISA K922830, Inova Quanta Lite SS-B ELISA K922832, Inova Quanta Lite Scl-70 ELISA K924898, Inova Quanta Lite Centromeres ELISA KOO3AEa, Inova Quanta Lite Jo-1 ELISA K922832, Inova Quanta Lite Ribosomal P ELISA K981237

Reference Device(s)

Not Found

Predetermined Change Control Plan (PCCP) - All Relevant Information

Not Found

Regulation Number and Section

§ 866.5100 Antinuclear antibody immunological test system.

(a)
Identification. An antinuclear antibody immunological test system is a device that consists of the reagents used to measure by immunochemical techniques the autoimmune antibodies in serum, other body fluids, and tissues that react with cellular nuclear constituents (molecules present in the nucleus of a cell, such as ribonucleic acid, deoxyribonucleic acid, or nuclear proteins). The measurements aid in the diagnosis of systemic lupus erythematosus (a multisystem autoimmune disease in which antibodies attack the victim's own tissues), hepatitis (a liver disease), rheumatoid arthritis, Sjögren's syndrome (arthritis with inflammation of the eye, eyelid, and salivary glands), and systemic sclerosis (chronic hardening and shrinking of many body tissues).(b)
Classification. Class II (performance standards).

Summary

0

K 1.23261

JUN 1 2 2013

ATTACHMENT I

PREMARKET NOTIFICATION 210(K) SAFETY AND EFFECTIVENESS SUMMARY (as required by 21 CFR § 807.92)

• A. 510(k)Number: K123261
• B. Purpose for Submission: New device
• C. Measurand:

D.

H.

• Anti-nRNP/Sm autoantibodies
• Type of Test: Qualitative enzyme immunoassay
• E. Applicant:
  • EUROIMMUN US INC.
• Proprietary and Established Names: ಿ. EUROIMMUN Anti-nRNP/Sm ELISA (IgG)
• G. Requlatory Information:
  • Regulation: 1.
    • 21 CFR 866.5110 Antinuclear antibody immunological test system
  • 2. Classification: Class II
  • 3. Product code: LKO
  • 4. Panel:

Immunology Intended Use:

• Intended use(s): 1 .
  The EUROIMMUN Anti-nRNP/Sm ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against nRNP/Sm in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of mixed connective tissue diseases and systemic lupus erythematosus, in conjunction with other laboratory and clinical findings.

• 2. Indication(s) for use: Same as intended use.
• 3. Special conditions for the use statement(s):

For prescription use only.

• 4. Special instrument requirements:
     Microwell plate reader capable of measuring OD at 450nm for dual wavelength readings.

l. Device Description:

The EUROIMMUN Anti-nRNP/Sm ELISA (IgG) consists of a microwell ELISA plate coated with nRNP/Sm antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution.

J. Substantial Equivalence Information:

• Predicate device name (s): 1 .
  • Inova Quanta Lite RNP ELISA
• 2. Predicate 510(k) number(s): K922833

1

3. Comparison with predicate:

Item	New Device	Predicate Device
Intended use	Detection of IgG antibodies to nRNP/Sm	Same
Technology	ELISA	Same
Assay platform	96-well microtiter plates	Same
Calibration	Relative evaluation	Same
Antigen	Purified U1-nRNP complex; U1-nRNP contains RNP as
well as Sm reactive proteins	Same
Conjugate	Anti-human IgG labeled with horseradish peroxidase	Same
Substrate	TMB	Same
Reagent preparation	All reagents, calibrators and controls are ready to use,
except for the wash buffer.	Same
Procedure	Sample incubation with micro-well antigen coated plate,
followed by a wash step, incubation with an anti-human
IgG enzyme conjugate; wash step, incubation with
substrate; then the addition of a stop solution and
reading at 450nm.	Same
Item	New Device	Predicate Device
Assay format	Qualitative	"Semi-quantitative" (using low positive control)
Sample dilution	1:201	1:101
Calibrators and
controls	1 calibrator
2 controls: 1 positive, 1 negative	3 controls: 1 high positive, 1 low positive (used for
calculation of results), 1 negative
Wash buffer	10x concentrate	40x concentrate
Stop solution	0.5 M sulphuric acid	0.344 M sulphuric acid
Sample types	Serum or plasma (EDTA, Li-heparin, Citrate)	Serum
Reported results	Ratio	Units
Cut off level	Ratio 1.0	20 Units

K. Standard/Guidance Document Referenced (if applicable):

Guidance for Industry and FDA Staff: Recommendations for Antibody (ANA) Test System Premarket (510(k)) Submissions (January 22, 2009)

L. Test Principle:

Patient samples are diluted 1:201 in sample buffer, 100 µl of each diluted patient sample and pre-diluted controls and calibrators are added to the antigen coated microtiter wells and incubated for 30 minutes at room temperature. After incubation the microtiter well strips are washed with wash buffer to remove unbound antibodies and 100 µl of the anti-human IgG enzyme conjugate reagent is added to each microtiter well. After an additional 30-minutes incubation at room temperature, the microtiter wells are again washed 3 times with 300 pl of wash buffer to remove any unbound enzyme conjugate and 100 µl of the chromogen substrate is added. The strips are incubated for 15 minutes at room temperature and 100 µl stop solution is added. The microtiter plates are placed in an ELISA reader and read at a wavelength of 450 nm and a reference wavelength of between 620 nm and 650 nm within 30 minutes.

Performance Characteristics (where applicable): M.

• Analytical performance:
  • Precision/Reproducibility: a.

The reproducibility of the test was investigated using sera with different concentrations. Intra-assay reproducibility is based on 20 determinations and inter-assay reproducibility on 40 determinations performed in 10 different runs on 5 days with 2 runs per day according to the package insert. The following results were obtained:

2

Intra-assay reproducibility

n = 20	Anti-nRNP/Sm ELISA (IgG) Ratio
	Sample 1	Sample 2	Sample 3	Sample 4	Sample 5	Sample 6
Mean value (x):	5.1	3.3	1.9	1.2	0.8	0.4
Range of values:	5.0 - 5.1	3.2 - 3.4	1.8 - 2.0	1.1 - 1.2	0.7 - 0.9	0.3 - 0.4
Expected result:	positive	positive	positive	positive	negative	negative
% positive:	100%	100%	100%	100%	0%	0%
% negative:	0%	0%	0%	0%	100%	100%

Inter-assay reproducibility

	Anti-nRNP/Sm ELISA (IgG)
n = 10 x 4	Ratio
	Sample 1	Sample 2	Sample 3	Sample 4	Sample 5	Sample 6
Mean value (x):	5.2	3.3	1.8	1.1	0.8	0.4
Range of values:	4.8 - 5.5	3.1 - 3.6	1.6 - 1.9	1.1 - 1.2	0.7 - 0.9	0.3 - 0.4
Expected result:	positive	positive	positive	positive	negative	negative
% positive:	100%	100%	100%	100%	0%	0%
% negative:	0%	0%	0%	0%	100%	100%

The lot to lot reproducibility was investigated during the validaton and quality control of the kit using different lots with QC samples distributed over the measurement range. The following results were obtained:

Lot to lot reproducibility

| | Anti-nRNP/Sm ELISA (IgG)
Ratio | | | | | | | Anti-nRNP/Sm ELISA (IgG)
Ratio | | | | |
|------------------|-----------------------------------|-----------|-----------|-----------|-----------|-----------|------------------|-----------------------------------|-----------|-----------|-----------|-----------|
| | Sample 1 | Sample 2 | Sample 3 | Sample 4 | Sample 5 | Sample 6 | | Sample 7 | Sample 8 | Sample 9 | Sample 10 | Sample 11 |
| n | 6* | 6* | 6* | 6* | 6* | 6* | n | 11** | 11** | 11** | 11** | 11** |
| Mean value (x): | 3.6 | 2.5 | 5.4 | 0.4 | 0.9 | 1.1 | Mean value (x): | 0.2 | 3.4 | 5.2 | 6.6 | 8.5 |
| Range of values: | 3.4 - 3.9 | 2.3 – 2.8 | 4.8 - 5.7 | 0.3 - 0.4 | 0.9 - 0.9 | 1.1 - 1.1 | Range of values: | 0.1 - 0.3 | 3.1 - 3.8 | 4.7 - 5.9 | 5.7 - 7.2 | 8.0 - 9.7 |
| Expected result: | positive | positive | positive | negative | negative | positive | Expected result: | negative | positive | positive | positive | positive |
| % positive: | 100% | 100% | 100% | 0% | 0% | 100% | % positive: | 0% | 100% | 100% | 100% | 100% |
| % negative: | 0% | 0% | 0% | 100% | 100% | 0% | % negative: | 100% | 0% | 0% | 0% | 0% |

*3 lots x 2 runs ** n lots x 1 run

b. Linearity/assay reportable range: Not applicable.

Not applicable.
High dose/leak-off.

C. High dose Hook effect

Potential for a high dose Hook effect is a phenomenon that is inherent with one step "sandwich" assay designs: Very high concentrations of antigen in the patient sample bind to all available sites saturating them - on both the antibody-solid phase and the antibody-labeled conjugate and thereby prevent the "sandwich" formation. Under these conditions, the measured level of analyte may be significantly lower than the actual level present in the sample. The two-step immunoassay design of the Anti-nRNP/Sm ELISA (IgG) eliminates the adverse contribution of binding proteins, endogenous interfering substances and general matrix effects due to the extra wash step.

3

Image /page/3/Picture/1 description: The image shows a pattern of alternating black and white shapes. The black shapes are arranged in a row and are separated by white spaces. The black shapes are rounded and symmetrical, resembling beads or capsules. The white spaces between the black shapes are rectangular and uniform in size.

d. Traceability, Stability, Expected values (controls, calibrators or methods):

A recognized standard or reference material for anti-nuclear antibodies is not available. Results of this assay are given in ratios. The reactivity of the Anti-nRNP/Sm ELISA was verified using the ANA reference panel of the CDC Centers for Disease Control and Prevention, Atlanta, USA.

• Limit of detection: e.
• Not applicable.
• f Analytical specificity:

Cross-reactivity: The quality of the antigen coated on the plates ensures a high specificity of the ELISA. Cross reactivity was investigated using a panel of 30 sera serologically positive for antibodies against rib.P-P, SS-A, SS-B, Scl-70, Jo-1, centromeres, Rubella virus, Measles virus, Herpes simplex virus type 1 and Borrelia burgdorferi. All 30 sera were negative in the Anti-nRNP/Sm ELISA (IgG), so no cross reactivity is expected.

Interference: To investigate the influence from hemoglobin, triglycerides and bilirubin, 5 different sera at different anti-nRNP/Sm concentrations (ratio 0.9 - 5.6) were spiked with potential interfering substances and were incubated with the test system acording to the package insert. Interferences from rheumatoid factor were investigated similarly by spiking with a RF positive serum. The recovery in relation to the unspiked sample without interferent was calculated. The individual recovery was within the range of 85 - 109 %. No significant interference was observed for concentrations of up to 1000 mg/dl for hemoglobin, 2000 mg/dl for triglyceride, 40 mg/dl for billrubin and 500 IU/ml for rheumatoid factor.

• Assay cut-off: g.
  Ratio 1.0

2. Comparison studies:

• Method comparison with predicate device: a.
  A comparison study was performed using 287 clinically characterized samples (52 MCTD, 69 SLE, 51 Sjögren's syndrome, 15 systemic sclerosis, 15 fibromyalgia, 35 RA, 30 borreliosis and 20 healthy), obtained from different sources from Europe and North America. The panel consisted of 49 men and 238 women. Age ranged from 14 to 85 years with an average age of 46 years. AntinRNP/Sm antibodies are expected in either MCTD or SLE. The other disease groups serve as control cohorts. The samples were tested with the EUROIMMUN Anti-nRNP/Sm ELISA (IgG) and with the Inova Quanta Lite RNP ELISA as the predicate device. Of the 8 discrepant samples, one was from a MCTD patient and the other 7 were from controls.

| All samples

n = 287		Predicate ELISA
		positive	negative
EUROIMMUN
Anti-nRNP/Sm ELISA (IgG)	positive	83	0
	negative	8	196
Negative agreement	$196 / 196 =$	100.0%	95% C.I.: 98.1% - 100.0%
Positive agreement	$83 / 91 =$	91.2%	95% C.I.: 83.4% - 96.1%
Overall agreement	$279 / 287 =$	97.2%	95% C.I.: 94.6% - 98.8%

• b. Matrix comparison:
  The usability of plasma was investigated using sample pairs each of serum and corresponding plasma (EDTA, Li-heparin, Citrate). Passing-Bablok regression was calculated for the comparison of serum to plasma. The regression equation is near the ideal correlation (intercept 0; slope 1.0) indicating equivalence of concentrations between serum and the corresponding plasma matrices. The comparison below satisfies this condition. Coefficients of determination were found to be above 0.99 and %recovery compared toserum was in the range of 91 to 120 % (serum = 100 %)

	EDTA plasma	Li-heparin plasma	Citrate plasma
n	15	15	15
Regression equation
(y = plasma, x = serum)	$y = 0.46 + 0.98 x$	$y = -0.25 + 1.05 x$	$y = 0.37 + 1.04$
95% C.I. of intercept	-0.56 - 2.41	-1.54 - 1.27	-1.72 - 1.82
95% C.I. of slope	0.95 - 1.01	0.97 - 1.09	0.97 - 1.09
Coefficient of determination R²	0.9988	0.9929	0.9915
Mean %recovery	101 %	103 %	105 %
Range of %recovery	91 - 117 %	92 - 114 %	93 - 120 %

4

Clinical studies:

Clinical studies were performed in cooperation with different sites. In total 1046 clinically characterized samples (65 from MCTD patients, 404 from SLE patients, 151 from myositis patients and 426 from control groups) were investigated for antibodies (IgG). With the EUROIMMUN Anti-nRNP/Sm ELISA (IgG) a prevalence of 100.0% (95% C.I.: 94.5 - 100.0%) was found in MCTD (clinical sensitivity) and a prevalence of 23.3% (95% C.I.: 19.2 - 27.7%) in SLE with a specificity of 99.3% (95% C.I.: 98.0 - 99.9%). The myositis panel was not considered for calculation of sensitivity and specificity, because anti-nRNP/Sm antibodies may occur in this disease (Tomer 1993). The results are shown in the table below. 95% C.I. are calculated by the exact method.

• Clinical sensitivity: a.

No.	Panel	n	Anti-nRNP/Sm ELISA (IgG)
			positive	%	95% C.I.
1	Mixed connective tissue diseases	65	65	100.0%	94.5 - 100.0%
2	Systemic lupus erythematosus	404	94	23.3%	19.2 - 27.7%

• Clinical specificity: b.

No.	Panel	n	Anti-nRNP/Sm ELISA (IgG)
			negative	%	95% C.I.
3	Polymyositis/dermatomyositis	151	143	94.7%	89.8 - 97.7%
4	Rheumatoid arthritis	164	164	100.0%	97.8 - 100.0%
5	Systemic sclerosis	81	81	100.0%	95.5 - 100.0%
6	Sjögren's syndrome	88	86	97.7%	92.0 - 99.7%
7	Other autoimmune diseases*	63	62	98.4%	91.5 - 100.0%
8	Borreliosis	30	30	100.0%	88.4 - 100.0%
	Total	577	566	98.1%	96.6 - 99.9%

*from the following groups: AIH (n = 8), Grave's disease (n = 12), Hashimoto (n = 11), celiac disease (n = 11), Diabetes Type I (n = 12)

• Other clinical supportive data (when a. and b. are not applicable): C. Not applicable.
• 4. Clinical cut-off:

See Assay cut-off.

• ട. Expected values/Reference range:
  The levels of anti-nRNP/Sm antibodies (IgG) were analyzed in a panel of 200 samples from apparently healthy blood donors (120 men and 80 women with an average age of 40 y; age range: 19 - 68 y). The results are shown in the table below.

n	200
Positives	1
Negatives	199
Prevalence	0.5%
	Ratio
Lowest value	0.1
Highest value	1.3
Mean value	0.1
Std deviation	0.09

N. Proposed Labeling:

The labeling is sufficient and it satisfies the requirements of 21 CFR Part 809.10.

O. Conclusion:

The submitted information in this premarket notification is complete and supports a substantial equivalence decision.

Michael Locke
Signature

Dir. Regulatory Affairs Title

12 June 2013 Date

5

ATTACHMENT 1

PREMARKET NOTIFICATION રાજીન્દ) SAFETY AND EFFECTIVENESS SUMMARY (as required by 21 CFR § 807.92)

• A. 510(k)Number: K123261
• B. Purpose for Submission: New device
• C. Measurand:
  • Anti-Sm autoantibodies
• D. Type of Test: Qualitative enzyme immunoassay
• E. Applicant:
  • EUROIMMUN US INC.
• F. Proprietary and Established Names: EUROIMMUN Anti-Sm ELISA (IgG)

G. Regulatory Information:

• 1. Regulation:
  • 21 CFR 866.5110 Antinuclear antibody immunological test system
• 2. Classification: Class II
• 3. Product code: LKP
• 4. Panel:

H.

• Immunology Intended Use:
• 1. Intended use(s):

The EUROIMMUN Anti-Sm ELISA (IgG) test kit is intended for the qualitative of IgG class autoantibodies against Sm in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of systemic lupus erythematosus, in conjunction with other laboratory and clinical findings.

• 2. Indication(s) for use:
  • Same as intended use.
• 3. Special conditions for the use statement(s):
  • For prescription use only.
• র্ম Special instrument requirements:

Microwell plate reader capable of measuring OD at 450nm for dual wavelength readings.

l. Device Description:

The EUROIMMUN Anti-Sm ELISA (IgG) consists of a microwell ELISA plate coated with Sm antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution.

J. Substantial Equivalence Information:

• Predicate device name (s): 1.
• Inova Quanta Lite Sm ELISA 2. Predicate 510(k) number(s):
• K922831

6

Image /page/6/Picture/1 description: The image shows a black and white illustration of a ladder. The ladder has four rungs that are evenly spaced apart. The rungs are connected to two vertical rails on either side.

3. Comparison with predicate:

Similarities
Item	New Device	Predicate Device
Intended use	Detection of IgG antibodies to Sm	Same
Technology	ELISA	Same
Assay platform	96-well microtiter plates	Same
Calibration	Relative units	Same
Antigen	Purified Sm antigen	Same
Conjugate	Anti-human IgG labeled with horseradish peroxidase	Same
Substrate	TMB	Same
Reagent preparation	All reagents, calibrator and controls are ready to use,
except for the wash buffer.	Same
Procedure	Sample incubation with micro-well antigen coated plate,
followed by a wash step, incubation with an anti-human
IgG enzyme conjugate; wash step, incubation with
substrate; then the addition of a stop solution and
reading at 450nm.	Same
Differences
Item	New Device	Predicate Device
Assay format	Qualitative	"Semi-quantitative" (using low positive control)
Sample dilution	1:201	1:101
Calibrators and	1 calibrator	3 controls: 1 high positive, 1 low positive (used for
controls	2 controls: 1 positive, 1 negative	calculation of results), 1 negative
Wash buffer	10x concentrate	40x concentrate
Stop solution	0.5 M sulphuric acid	0.344 M sulphuric acid
Sample types	Serum or plasma (EDTA, Li-heparin, Citrate)	Serum
Reported results	Ratio	Units
Cut off level	Ratio 1.0	20 Units

K. Standard/Guidance Document Referenced (if applicable):

Guidance for Industry and FDA Staff: Recommendations for Antibody (ANA) Test System Premarket (510(k)) Submissions (January 22, 2009)

L. Test Principle:

Patient samples are diluted 1:201 in sample buffer, 100 pl of each diluted patient sample and pre-diluted controls and calibrators are added to the antigen coated microtiter wells and incubated for 30 minutes at room temperature. After incubation the microtier well strips are washed with wash buffer to remove unbound antibodies and 100 ul of the anti-human IgG enzyme coniugate reagent is added to each microtiter well. After an additional 30-minutes incubation at room temperature, the microtiter wells are again washed 3 times with 300 µl of wash buffer to remove any unbound enzyme conjugate and 100 µl of the chromogen substrate is added. The strips are incubated for 15 minutes at room temperature and 100 ul stop solution is added. The microtiter plates are placed in an ELISA reader and read at a wavelength of 450 nm and a reference wavelength of between 620 nm and 650 nm within 30 minutes.

Performance Characteristics (where applicable): M.

• Analytical performance:
  • Precision/Reproducibility: a.

The reproducibility of the test was investigated using sera with different concentrations. Intra-assay reproducibility is based on 20 determinations and inter-assay reproducibility on 40 determinations performed in 10 different runs on 5 days with 2 runs per day according to the package insert. The following results were obtained:

7

Image /page/7/Picture/0 description: The image shows a repeating pattern of black shapes against a white background. The black shapes are vertically oriented and have a rounded, almost pill-like form, with a narrower section in the middle. These shapes are evenly spaced and aligned in a row, creating a rhythmic visual sequence. The top and bottom of the image are bordered by solid black lines, which frame the repeating pattern.

Intra-assay reproducibility Anti-Sm ELISA (IgG) Ratio n = 20 Sample 2 Sample 5 Sample 6 Sample 1 Sample 3 Sample 4 Mean value (x): 5.7 3.7 2.0 1.2 0.8 0.4 5.4 - 5.8 3.5 - 3.9 1.9 – 2.0 1.2 - 1.3 0.7 - 0.9 0.4 - 0.4 Range of values: negative Expected result: positive positive positive positive negative % positive: 100% 100% 100% 100% 0% 0% 0% 100% 100% % negative: 0% 0% 0%

Inter-assay reproducibility

| n = 10 x 4 | Anti-Sm ELISA (IgG)
Ratio | | | | | |
|------------------|------------------------------|-----------|-----------|-----------|-----------|-----------|
| | Sample 1 | Sample 2 | Sample 3 | Sample 4 | Sample 5 | Sample 6 |
| Mean value (x): | 5.8 | 3.9 | 1.7 | 1.1 | 0.8 | 0.4 |
| Range of values: | 5.3 - 7.4 | 3.5 - 4.6 | 1.5 - 2.2 | 1.0 - 1.2 | 0.6 - 1.0 | 0.3 - 0.5 |
| Expected result: | positive | positive | positive | positive | negative | negative |
| % positive: | 100% | 100% | 100% | 100% | 2.5% | 0% |
| % negative: | 0% | 0% | 0% | 0% | 97.5% | 100% |

The lot to lot reproducibility was investigated during the validaton and quality control of the kit using different lots with QC samples distributed over the measurement range. The following results were obtained:

Lot to lot reproducibility

		Anti-Sm ELISA (IgG)
		Ratio
	Sample 1	Sample 2	Sample 3	Sample 4	Sample 5	Sample 6
n	6*	6*	6*	6*	6*	6*
Mean value (x):	3.6	3.4	3.1	0.3	0.9	1.2
Range of values:	3.3 - 3.9	3.2 - 3.9	2.9 - 3.4	0.3 - 0.3	0.8 - 0.9	1.1 - 1.3
Expected result:	positive	positive	positive	negative	negative	positive
% positive:	100%	100%	100%	0%	0%	100%
% negative:	0%	0%	0%	100%	100%	0%

	Anti-Sm ELISA (IgG) Ratio
	Sample 7	Sample 8	Sample 9	Sample 10	Sample 11
n	9**	10**	10**	9**	10**
Mean value (x):	0.1	1.9	3.4	5.7	8.3
Range of values:	0.0 - 0.2	1.5 - 2.2	2.6 - 3.8	4.9 - 6.7	7.5 - 9.6
Expected result:	negative	positive	positive	positive	positive
% positive:	0%	100%	100%	100%	100%
% negative:	100%	0%	0%	0%	0%

*3 lots x 2 runs ** n lots x 1 run

Linearity/assay reportable range: b. Not applicable.

High dose Hook effect C.

Potential for a high dose Hook effect is a phenomenon that is inherent with one step "sandwich" assay designs: Very high concentrations of antigen in the patient sample bind to all available sites saturating them - on both the antibody-solid phase and the antibody-labeled conjugate and thereby prevent the "sandwich" formation. Under these conditions, the measured level of analyte may besignificantly lower than the actual level present in the sample. The two-step immunoassay design of the Anti-Sm ELISA (IgG) eliminates the adverse contribution of binding proteins, endogenous interfering substances and general matrix effects due to the extra wash step.

8

Image /page/8/Picture/1 description: The image shows a black and white pattern. There are four black shapes that are evenly spaced. The shapes are connected to a black line at the bottom of the image. There is a white line at the top and bottom of the image.

d. Traceability, Stability, Expected values (controls, calibrators or methods):

A recognized standard or reference material for anti-nuclear antibodies is not available. Results of this assay are given in ratios. The reactivity of the Anti-Sm ELISA was verified using the ANA reference panel of the CDC Centers for Disease Control and Prevention, Atlanta, USA.

• Limit of detection: e.
• Not applicable
• f. Analvtical specificity:

Cross-reactivity: The quality of the antigen coated on the plates ensures a high specificity of the ELISA. Cross reactivity was investigated using a panel of 30 sera serologically positive for antibodies against rib.P-P, nRNP/Sm, SS-A, Scl-70, Jo-1, centromeres, Rubella virus, Measles virus, Herpes simplex virus type 1 and Borrelia burgdorferi. All 30 sera were negative in the Anti-Sm ELISA (IgG), so no cross reactivity is expected.

Interference: To investigate the influence from hemoglobin, triglycerides and bilirubin, 5 different sera at different anti-Sm concentrations (ratio 0.7 ~ 3.9) were spiked with potential interfering substances and were incubated with the test system according to the package insert. Interferences from rheumatoid factor were investigated similarly by spiking with a RF positive serum. The recovery in relation to the unspiked sample without interferent was calculated. The individual recovery was within the range of 90 - 111 %. No significant interference was observed for concentrations of up to 1000 ma/dl for hemoalobin, 2000 mg/dl for trialyceride, 40 mg/dl for billrubin and 500 lU/ml for rheumatoid factor.

• Assay cut-off: ਉਂ
• Ratio 1.0
• 2. Comparison studies:
  • Method comparison with predicate device: a.

A comparison study was performed using 294 clinically characterized samples (128 SLE, 51 Sjögren's syndrome, 15 systemic sclerosis, 15 fibromyalgia, 35 RA, 30 borreliosis and 20 healthy), obtained from different sources from Europe and North America. The panel consisted of 60 men and 234 women. Age ranged from 14 to 85 years with an average age of 45 years. Anti-Sm antibodies are expected in SLE. The other disease groups serve as control cohorts. The samples were tested with the EUROMMUN Anti-Sm ELISA (IqG) and with the Inova Quanta Lite Sm ELISA as the predicate device. The results are shown in the table below. All of the 7 discrepant samples negative in the EUROIMMUN test were from controls. The 5 discrepant samples positive in the EUROIMMUN test were from SLE patients.

| All samples

n = 294		Predicate ELISA
		positive	negative
EUROIMMUN
Anti-Sm ELISA (IgG)	positive	37	5
	negative	7	245
Negative agreement	$245 / 250 = 98.0%$	95% C.I.: 95.4%	99.3%
Positive agreement	$37 / 44 = 84.1%$	95% C.I.: 69.9%	93.4%
Overall agreement	$282 / 294 = 95.9%$	95% C.I.: 93.0%	97.9%

• b. Matrix comparison:
  The usability of plasma was investigated using sample pairs each of serum and corresponding plasma (EDTA, Li-heparin, Citrate). Passing-Bablok regression was calculated for the comparison of serum to plasma. The regression equation is near the ideal correlation (intercept 0) indicating equivalence of concentrations between serum and the corresponding plasma matrices. Coefficients of determination were found to be above 0.99 and %recovery compared to serum was in the range of 89 to 118 % (serum = 100 %).

	EDTA plasma	Li-heparin plasma	Citrate plasma
n	16	16	16
Regression equation
(y = plasma, x = serum)	$y = 0.33 + 1.00x$	$y = 0.19 + 0.97x$	$y = 0.05 + 1.02$
95% C.I. of intercept	-0.32 - 1.24	-0.92 - 1.95	-0.69 - 3.27
95% C.I. of slope	0.96 - 1.03	0.94 - 1.01	0.99 - 1.05
Coefficient of determination R2	0.9953	0.9948	0.9931
Mean %recovery	102 %	99 %	103 %
Range of %recovery	94 - 115 %	90 - 111 %	89 - 118 %

9

3. Clinical studies:

Clinical studies were performed in cooperation with different sites. In total 1036 clinically characterized samples (414 from SLE patients and 622 from control groups) were investigated for antibodies (IgG). With the EUROIMMUN Anti-Sm ELISA (IgG) a prevalence of 11.4% (95% C.I.: 8.5 - 14.8%) was found in SLE with a specificity of 99.0% (95% C.I.: 97.9 - 99.6%). The results are shown in the table below. 95% C.I. are calculated by the exact method.

• a. Clinical sensitivity:

No.	Panel	n	Anti-Sm ELISA (IgG)
			positive	%	95% C.I.
1	Systemic lupus erythematosus	414	47	11.4%	8.5 - 14.8%
No.	Panel	n	negative	%	95% C.I.
2	Rheumatoid arthritis	164	164	100.0%	97.8 - 100.0%
3	Systemic sclerosis	81	81	100.0%	95.5 - 100.0%
4	Sjögren's syndrome	88	88	100.0%	95.9 - 100.0%
5	Polymyositis/dermatomyositis	151	151	100.0%	97.6 - 100.0%
6	Mixed connective tissue diseases	45	39	86.7%	73.2 - 94.9%
7	Other autoimmune diseases*	63	63	100.0%	94.3 - 100.0%
8	Borreliosis	30	30	100.0%	88.4 - 100.0%
	Total	622	616	99.0%	97.9 - 99.6%

b. Clinical specificity:

*from the following groups: AIH (n = 8), Grave's disease (n = 12), Hashimoto (n = 11), celiac disease (n = 11), Diabetes Type I (n = 12)

• Other clinical supportive data (when a. and b. are not applicable): C. Not applicable.
• 4. Clinical cut-off:

See Assay cut-off.

• ട. Expected values/Reference range:
  The levels of anti-Sm antibodies (IgG) were analyzed in a panel of 200 samples from apparently healthy blood donors (120 men and 80 women with an average age of 40 y; age range: 19 - 68 y). The results are shown in the table below.

n	200
Positives	0
Negatives	200
Prevalence	0.0%
	Ratio
Lowest value	0.1
Highest value	0.3
Mean value	0.1
Std deviation	0.02

N. Proposed Labeling:

The labeling is sufficient and it satisfies the requirements of 21 CFR Part 809.10.

0. Conclusion:

The submitted information in this premarket notification is complete and supports a substantial equivalence. decision.

Michael Locke
Signature

Dir. Regulatory Affairs

12 June 2013 Date

10

Image /page/10/Picture/1 description: The image shows a series of four dark, circular shapes positioned between two horizontal lines. The circular shapes are evenly spaced and appear to be touching both the top and bottom lines. The overall composition is simple and graphic, with a focus on the contrast between the dark shapes and the surrounding space.

ATTACHMENT 1

PREMARKET NOTIFICATION 510(K) SAFETY AND EFFECTIVENESS SUMMARY (as required by 21 CFR § 807.92)

• A. 510(k)Number: K123261
• Purpose for Submission: B. New device
• ். Measurand:
  • Anti-SS-A autoantibodies ·
• D. Type of Test:
• Qualitative enzyme immunoassay ய் Applicant:
  • EUROIMMUN US INC.
• Proprietary and Established Names: F. EUROIMMUN Anti-SS-A ELISA (IgG)

G. Regulatory Information:

• Regulation: 1.
  • 21 CFR 866.5100 Antinuclear antibody immunological test system
• 2. Classification:
• Class II
• 3. Product code:
• LLL
• 4. Panel:
• Immunology H. Intended Use:
  • Intended use(s): 1.

The EUROIMMUN Anti-SS-A ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against SS-A in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of Sjögren's syndrome and systemic lupus erythematosus, in conjunction with other laboratory and clinical findings.

• 2. Indication(s) for use: Same as intended use.
• 3. Special conditions for the use statement(s): For prescription use only.
• র্ব : Special instrument requirements:

Microwell plate reader capable of measuring OD at 450nm for dual wavelength readings. Device Description:

l:

The EUROIMMUN Anti-SS-A ELISA (IgG) consists of a microwell ELISA plate coated with SS-A antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate. TMB chromogen/substrate solution and stop solution.

Substantial Equivalence Information: J.

• Predicate device name (s): 1.
  • Inova Quanta Lite SS-A ELISA
• 2. Predicate 510(k) number(s): K922830

11

Image /page/11/Picture/1 description: The image shows the number 3 followed by the text 'Comparison with predicate:'. The number 3 is on the left side of the image. The text is on the right side of the image. The text is underlined.

Similarities
Item	New Device	Predicate Device
Intended use	Detection of IgG antibodies to SS-A	Same
Technology	ELISA	Same
Assay platform	96-well microtiter plates	Same
Calibration	Relative units	Same
Antigen	Purified SS-A antigen	Same
Conjugate	Anti-human IgG labeled with horseradish peroxidase	Same
Substrate	TMB	Same
Reagent preparation	All reagents, calibrator and controls are ready to use,
except for the wash buffer.	Same
Procedure	Sample incubation with micro-well antigen coated plate,
followed by a wash step, incubation with an anti-human
IgG enzyme conjugate; wash step, incubation with
substrate; then the addition of a stop solution and
reading at 450nm.	Same
Differences
Item	New Device	Predicate Device
Assay format	Qualitative	"Semi-quantitative" (using low positive control)
Sample dilution	1:201	1:101
Calibrators and	1 calibrator	3 controls: 1 high positive, 1 low positive (used for
controls	2 controls: 1 positive, 1 negative	calculation of results), 1 negative
Wash buffer	10x concentrate	40x concentrate
Stop solution	0.5 M sulphuric acid	0.344 M sulphuric acid
Sample types	Serum or plasma (EDTA, Li-heparin, Citrate)	Serum
Reported results	Ratio	Units
Cut off level	Ratio 1.0	20 Units

Standard/Guidance Document Referenced (if applicable): K.

Guidance for Industry and FDA Staff: Recommendations for Antibody (ANA) Test System Premarket (510(k)) Submissions (January 22, 2009)

ட. Test Principle:

Patient samples are diluted 1:201 in sample buffer, 100 µl of each diluted patient sample and pre-diluted controls and calibrators are added to the antigen coated microtiter wells and incubated for 30 minutes at room temperature. After incubation the microtiter well strips are washed with wash buffer to remove unbound antibodies and 100 µl of the anti-human IgG enzyme conjugate reagent is added to each microtiter well. After an additional 30-minutes incubation at room temperature, the microtiter wells are again washed 3 times with 300 ul of wash buffer to remove any unbound enzyme conjugate and 100 µl of the chromogen substrate is added. The strips are incubated for 15 minutes at room temperature and 100 µl stop solution is added. The microtiter plates are placed in an ELISA reader and read at a wavelength of 450 nm and a reference wavelength of between 620 nm and 650 nm within 30 minutes.

Performance Characteristics (where applicable): M.

• Analytical performance:
• Precision/Reproducibility: ·
  • The reproducibility of the test was investigated using sera with different concentrations. Intra-assay reproducibility is based on 20 determinations and inter-assay reproducibility on 40 determinations performed in 10 different runs on 5 days with 2 runs per day, each run performed according to the package insert. The following results were obtained:

Image /page/11/Picture/13 description: The image shows a pattern of alternating black and white shapes. The black shapes are rectangular bars, and the white shapes are oval-like shapes. The black bars are arranged horizontally, and the white shapes are arranged vertically between the bars. There are four white shapes in the image.

12

Image /page/12/Picture/0 description: The image shows a pattern of black shapes against a white background. The shapes are arranged in a row, with each shape consisting of two horizontal lines connected by a rounded, oval-like form in the middle. The pattern is symmetrical and evenly spaced, creating a repeating design.

Intra-assay reproducibility

| n = 20 | Anti-SS-A ELISA (IgG)
Ratio | | | | | |
|------------------|--------------------------------|-----------|-----------|-----------|-----------|-----------|
| | Sample 1 | Sample 2 | Sample 3 | Sample 4 | Sample 5 | Sample 6 |
| Mean value (x): | 6.1 | 4.2 | 2.0 | 1.3 | 0.8 | 0.3 |
| Range of values: | 5.9 - 6.4 | 4.0 - 4.5 | 1.9 - 2.1 | 1.2 - 1.4 | 0.7 - 0.8 | 0.3 - 0.3 |
| Expected result: | positive | positive | positive | positive | negative | negative |
| % positive: | 100% | 100% | 100% | 100% | 0% | 0% |
| % negative: | 0% | 0% | 0% | 0% | 100% | 100% |

Inter-assay reproducibility

| n = 10 x 4 | Anti-SS-A ELISA (IgG)
Ratio | | | | | |
|------------------|--------------------------------|-----------|-----------|-----------|-----------|-----------|
| | Sample 1 | Sample 2 | Sample 3 | Sample 4 | Sample 5 | Sample 6 |
| Mean value (x): | 6.0 | 4.0 | 1.8 | 1.2 | 0.8 | 0.3 |
| Range of values: | 5.0 - 6.4 | 3.7 - 4.5 | 1.6 - 1.9 | 1.0 - 1.4 | 0.7 - 0.9 | 0.3 - 0.4 |
| Expected result: | positive | positive | positive | positive | negative | negative |
| % positive: | 100% | 100% | 100% | 100% | 0% | 0% |
| % negative: | 0% | 0% | 0% | 0% | 100% | 100% |

The lot to lot reproducibility was investigated during the validaton and quality control of the kit using different lots with QC samples distributed over the measurement range. The following results were obtained:

Lot to lot reproducibility

| | Anti-SS-A ELISA (IgG)
Ratio | | | | | |
|------------------|--------------------------------|-----------|-----------|-----------|-----------|-----------|
| | Sample 1 | Sample 2 | Sample 3 | Sample 4 | Sample 5 | Sample 6 |
| n | 6* | 6* | 6* | 6* | 6* | 6* |
| Mean value (x): | 4.6 | 5.6 | 5.6 | 0.3 | 0.9 | 1.1 |
| Range of values: | 4.4 - 4.8 | 5.1 - 6.2 | 5.1 - 6.0 | 0.3 - 0.4 | 0.9 - 0.9 | 1.1 - 1.2 |
| Expected result: | positive | positive | positive | negative | negative | positive |
| % positive: | 100% | 100% | 100% | 0% | 0% | 100% |
| % negative: | 0% | 0% | 0% | 100% | 100% | 0% |

| | Anti-SS-A ELISA (IgG)
Ratio | | | | |
|------------------|--------------------------------|-----------|-----------|-----------|-----------|
| | Sample 7 | Sample 8 | Sample 9 | Sample 10 | Sample 11 |
| n | 10** | 11** | 10** | 9** | 11** |
| Mean value (x): | 0.1 | 1.6 | 2.7 | 4.6 | 7.8 |
| Range of values: | 0.1 - 0.2 | 1.3 - 1.8 | 2.3 - 3.1 | 4.3 - 5.2 | 7.4 - 8.3 |
| Expected result: | negative | positive | positive | positive | positive |
| % positive: | 0% | 100% | 100% | 100% | 100% |
| % negative: | 100% | 0% | 0% | 0% | 0% |

*3 lots x 2 runs ** n lots x 1 run

b. Linearity/assay reportable range: Not applicable.

Hiah dose Hook effect C.

Potential for a high dose Hook effect is a phenomenon that is inherent with one step "sandwich" assay designs: Very high concentrations of antigen in the patient sample bind to all available sites saturating them - on both the antibody-solid phase and the antibody-labeled conjugate and thereby prevent the "sandwich" formation. Under these conditions, the measured level of analyte may be significantly lower than the actual level present in the sample. The two-step immunoassay design of the Anti-SS-A ELISA (IgG) eliminates the adverse contribution of binding proteins, endogenous interfering substances and general matrix effects due to the extra wash step.

13

Image /page/13/Picture/1 description: The image shows a pattern of alternating black and white shapes. There are four black, oval-like shapes evenly spaced between two horizontal black lines. The white space between the black shapes and lines creates a contrasting pattern.

Traceability, Stability, Expected values (controls, calibrators or methods): d.

A recognized standard or reference material for anti-nuclear antibodies is not available. Results of this assay are given in ratios. The reactivity of the Anti-SS-A ELISA was verified using the ANA reference panel of the CDC Centers for Disease Control and Prevention, Atlanta, USA.

• e. Limit of detection:
• Not applicable.
• f. Analytical specificity:

Cross-reactivity: The quality of the antigen coated on the plates ensures a high specificity of the ELISA. Cross reactivity was investigated using a panel of 30 sera serologically positive for antibodies against rib.P-P, nRNP/Sm, Sm, Scl-70, Jo-1, centromeres, Rubella virus, Measles virus, Herpes simplex virus type 1 and Borrelia burgdorferi. All 30 sera were negative in the Anti-SS-A ELISA (IgG), so no cross reactivity is expected.

Interference: To investigate the influence from hemoglobin, triglycerides and bilirubin, 5 different sera at different anti-SS-A concentrations (15 - 124 RU/ml) were spiked with potential interfering substances and were incubated with the test system according to the package insert. Interferences from rheumatoid factor were investigated similarly by spiking with a RF positive serum. The recovery in relation to the unspiked sample without interferent was calculated. The individual recovery was within the range of 93 - 107 %. No significant interference was observed for concentrations of up to 1000 mg/dl for hemoglobin, 2000 mg/dl for triglyceride, 40 mg/dl for billirubin and 500 IU/ml for rheumatoid factor.

• ದ್ರ. Assay cut-off:
  . .

Ratio 1.0

2. Comparison studies:

• Method comparison with predicate device: a.
  A comparison study was performed using 305 clinically characterized samples (63 SLE, 77 Sjögren's syndrome, 23 systemic sclerosis, 1 SSc/SS, 15 fibromyalgia, 26 myositis, 35 RA, 30 borreliosis, 20 healthy), obtained from different sources from Europe and North America. The panel consisted of 54 men and 251 women. Age ranged from 14 to 85 years with an average age of 48 years. Anti-SS-A antibodies are expected in either Siögren's syndrome or SLE. The other disease groups serve as control cohorts. The samples were tested with the EUROIMMUN Anti-SS-A ELISA (IgG) and with the Inova Quanta Lite SS-A ELISA as the predicate device. Of the 9 discrepant samples negative in the EUROIMMUN test. 2 were from patients with Siögren's syndrome and one from a SLE patient, the other 6 were from controls. All 3 discrepant samples positive in the EUROIMMUN test were from SLE patients.

| All samples

n = 305		Predicate ELISA
		positive	negative
EUROIMMUN
Anti-SS-A ELISA (IgG)	positive	116	3
	negative	9	177
Negative agreement    177 / 180 = 98.3%      95% C.I.: 95.2% - 99.7%
Positive agreement    116 / 125 = 92.8%      95% C.I.: 86.8% - 96.7%
Overall agreement      293 / 305 = 96.1%      95% C.I.: 93.2% - 98.0%

• b. Matrix comparison:
  The usability of plasma was investigated using sample pairs each of serum and corresponding plasma (EDTA, Li-heparin, Citrate). Passing-Bablok regression was calculated for the comparison of serum to plasma. The regression equation is near the ideal correlation (intercept 0; slope 1.0) indicating equivalence of concentrations between serum and the corresponding plasma matrices. Coefficients of determination were found to be above 0.99 and %recovery compared to serum was in the range of 85 to 104 % (serum = 100 %).

	EDTA plasma	Li-heparin plasma	Citrate plasma
n	16	16	16
Regression equation
(y = plasma, x = serum)	$y = -1.59 + 0.99 x$	$y = -1.47 + 0.99 x$	$y = -0.54 + 0.98$
95% C.I. of intercept	-4.49 - 3.01	-4.62 - 1.36	-4.65 - 3.32
95% C.I. of slope	0.95 - 1.02	0.95 - 1.02	0.94 - 1.04
Coefficient of determination R²	0.9972	0.9929	0.9956
Mean %recovery	97 %	95 %	98 %
Range of %recovery	90 - 102 %	85 - 102 %	93 - 104 %

14

Clinical studies:

Clinical studies were performed in cooperation with different sites. In total 1026 clinically characterized samples (88 from SS patients, 404 from SLE patients and 534 from control groups) were investigated for anti-SS-A antibodies (IgG). With the EUROIMMUN Anti-SS-A ELISA (IgG) a prevalence of 73.9% (95% C.I.: 63.4 – 82.7%) was found in Sjögren's syndrome (clinical sensitivity) and a prevalence of 40.6% (95% C.I.: 35.8 - 45.6%) was found in SLE with a specificity of 94.8% (95% C.I.: 92.5 - 96.5%). Systemic sclerosis and myositis panels were considered for calculation of specificity, however, it has been reported that anti-SS-A antibodies may occur in these diseases [Antonioli 2002, Ghirardello 2005]. The results are shown in the table below. 95% C.I. are calculated by the exact method.

• Clinical sensitivity: a.

No.	Panel	n	positive	%	95% C.I.
1	Sjögren's syndrome	88	65	73.9%	63.4 - 82.7%
2	Systemic lupus erythematosus	404	164	40.6%	35.8 - 45.6%

Clinical specificity: b.

No.	Panel	n	negative	%	95% C.I.
3	Systemic sclerosis	81	75	92.6%	84.6 - 97.2%
4	Polymyositis/dermatomyositis	151	138	91.4%	85.7 - 95.3%
5	Rheumatoid arthritis	164	159	97.0%	93.0 - 99.0%
6	Mixed connective tissue diseases	45	41	91.1%	78.8 - 97.5%
7	Other autoimmune diseases*	63	63	100.0%	94.3 - 100.0%
8	Borreliosis	30	30	100.0%	88.4 - 100.0%
	Total	534	506	94.8%	92.5 - 96.5%

ffrom the following groups: AIH (n = 8), Grave's disease (n = 12), Hashimoto (n = 11), celiac disease (n = 11), Diabetes Type I (n = 12)

• Other clinical supportive data (when a. and b. are not applicable). C. Not applicable.
• 4. Clinical cut-off:

See Assay cut-off.

• 5. Expected values/Reference range:
     The levels of anti-SS-A antibodies (IgG) were analyzed in a panel of 200 samples from apparently healthy blood donors (120 men and 80 women with an average age of 40 y; age range: 19 - 68 y). The results are shown in the table below.

n	200
Positives	2
Negatives	198
Prevalence	1.0%
	Ratio
Lowest value	0.0
Highest value	4.4
Mean value	0.1
Std deviation	0.33

N. Proposed Labeling:

The labeling is sufficient and it satisfies the requirements of 21 CFR Part 809.10.

O. Conclusion:

The submitted information in this premarket notification is complete and supports a substantial equivalence decision.

Michael Locke

Signature

Dir. Regulatory Affairs Title

12 June 2013 Date

15

Image /page/15/Picture/1 description: The image shows a pattern of alternating black and white shapes. The black shapes are arranged in a row and are separated by white spaces. The black shapes are rounded and bulbous, while the white spaces are rectangular. The pattern is symmetrical and repeats several times across the image.

ATTACHMENT 1

PREMARKET NOTIFICATION 210(K) SAFETY AND EFFECTIVENESS SUMMARY (as required by 21 CFR § 807.92)

• A. 510(k)Number: K123261
• Purpose for Submission: в. New device
• C. Measurand:
  • Anti-SS-B autoantibodies
• D. Type of Test:
  • Qualitative enzyme immunoassay
• E. Applicant:
  • EUROIMMUN US INC.
• F. Proprietary and Established Names: EUROIMMUN Anti-SS-B ELISA (IqG)

Regulatory Information: G.

• 1. Regulation:
  • 21 CFR 866.5110 Antinuclear antibody immunological test system
• 2. Classification: Class II
• 3. Product code: LLL
• র্ব Panel:
• Immunology
• H. Intended Use:
  • Intended use(s): 1.

The EUROIMMUN Anti-SS-B ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against SS-B in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of Sjögren's syndrome and systemic lupus erythematosus, in conjunction with other laboratory and clinical findings.

• 2. Indication(s) for use: Same as intended use.
• ന Special conditions for the use statement(s): For prescription use only .

Special instrument requirements: 4.

Microwell plate reader capable of measuring OD at 450nm for dual wavelength readings. Device Description:

l.

The EUROIMMUN Anti-SS-B ELISA (IgG) consists of a microwell ELISA plate coated with SS-B antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution.

Substantial Equivalence Information: J.

• Predicate device name (s): 1.
  Inova Quanta Lite SS-B ELISA

• 2. Predicate 510(k) number(s): K922832

16

റ്റ് Comparison with predicate:

Similarities
Item	New Device	Predicate Device
Intended use	Detection of IgG antibodies to SS-B	Same
Technology	ELISA	Same
Assay platform	96-well microtiter plates	Same
Calibration	Relative units	Same
Antigen	Purified SS-B antigen	Same
Conjugate	Anti-human IgG labeled with horseradish peroxidase	Same
Substrate	TMB	Same
Reagent preparation	All reagents, calibrator and controls are ready to use,
except for the wash buffer.	Same
Procedure	Sample incubation with micro-well antigen coated plate,
followed by a wash step, incubation with an anti-human
IgG enzyme conjugate; wash step, incubation with
substrate; then the addition of a stop solution and
reading at 450nm.	Same
Differences
Item	New Device	Predicate Device
Assay format	Qualitative	"Semi-quantitative" (using low positive control)
Sample dilution	1:201	1:101
Calibrators and	1 calibrator	3 controls: 1 high positive, 1 low positive (used for
controls	2 controls: 1 positive, 1 negative	calculation of results), 1 negative
Wash buffer	10x concentrate	40x concentrate
Stop solution	0.5 M sulphuric acid	0.344 M sulphuric acid
Sample types	Serum or plasma (EDTA, Li-heparin, Citrate)	Serum
Reported results	Ratio	Units
Cut off level	Ratio 1.0	20 Units

Standard/Guidance Document Referenced (if applicable): K.

Guidance for Industry and FDA Staff: Recommendations for Antibody (ANA) Test System Premarket (510(k)) Submissions (January 22, 2009)

L. Test Principle:

Patient samples are diluted 1:201 in sample buffer, 100 pl of each diluted patient sample and pre-diluted controls and calibrators are added to the antigen coated microtiter wells and incubated for 30 minutes at room temperature. After incubation the microtiter well strips are washed with wash buffer to remove unbound antibodies and 100 ul of the anti-human IgG enzyme conjugate reagent is added to each microtiter well. After an additional 30-minutes incubation at room temperature, the microtiter wells are again washed 3 times with 300 ul of wash buffer to remove any unbound enzyme conjugate and 100 µl of the chromogen substrate is added. The strips are incubated for 15 minutes at room temperature and 100 ul stop solution is added. The microtiter plates are placed in an ELISA reader and read at a wavelength of 450 nm and a reference wavelength of between 620 nm and 650 nm within 30 minutes.

Performance Characteristics (where applicable): M.

• Analytical performance: 1 -
  • Precision/Reproducibility: a.

The reproducibility of the test was investigated using sera with different concentrations. Intra-assay reproducibility is based on 20 determinations and inter-assay reproducibility on 40 determinations performed in 10 different runs on 5 days with 2 runs per day, each run performed according to the package insert. The following results were obtained:

17

Image /page/17/Picture/0 description: The image shows a close-up of a black and white pattern. The pattern consists of a series of vertical black bars that are connected by horizontal black lines at the top and bottom. The spaces between the vertical bars are white and have a rounded, oval shape. There are four of these oval shapes visible in the image.

Intra-assay reproducibility
			Anti-SS-B ELISA (IgG)
n = 20			Ratio
	Sample 1	Sample 2	Sample 3	Sample 4	Sample 5	Sample 6
Mean value (x):	4.1	2.9	1.3	0.8	0.6	0.4
Range of values:	4.0 - 4.3	2.8 - 3.1	1.1 - 1.4	0.7 - 0.9	0.5 - 0.7	0.4 - 0.5
Expected result:	positive	positive	positive	negative	negative	negative
% positive:	100%	100%	100%	0%	0%	0%
% negative:	0%	0%	0%	100%	100%	100%

Inter-assay reproducibility

| n = 10 x 4 | Anti-SS-B ELISA (IgG)
Ratio | | | | | |
|------------------|--------------------------------|-----------|-----------|-----------|-----------|-----------|
| | Sample 1 | Sample 2 | Sample 3 | Sample 4 | Sample 5 | Sample 6 |
| Mean value (x): | 4.7 | 3.2 | 1.5 | 1.1 | 0.7 | 0.5 |
| Range of values: | 4.2 - 5.3 | 3.0 - 3.7 | 1.3 - 1.7 | 1.0 - 1.1 | 0.5 - 0.7 | 0.4 - 0.5 |
| Expected result: | positive | positive | positive | positive | negative | negative |
| % positive: | 100% | 100% | 100% | 100% | 0% | 0% |
| % negative: | 0% | 0% | 0% | 0% | 100% | 100% |

The lot to lot reproducibility was investigated during the validaton and quality control of the kit using different lots with QC samples distributed over the measurement range. The following results were obtained:

Lot to lot reproducibility

	Anti-SS-B ELISA (IgG) Ratio
	Sample 1	Sample 2	Sample 3	Sample 4	Sample 5	Sample 6
n	6*	6*	6*	6*	6*	6*
Mean value (x):	3.9	3.9	3.8	0.3	0.9	1.2
Range of values:	3.6 - 4.2	3.4 - 4.4	3.3 - 4.1	0.3 - 0.3	0.9 - 0.9	1.1 - 1.3
Expected result:	positive	positive	positive	negative	negative	positive
% positive:	100%	100%	100%	0%	0%	100%
% negative:	0%	0%	0%	100%	100%	0%

	Anti-SS-B ELISA (IgG) Ratio
	Sample 7	Sample 8	Sample 9
n	11**	11**	11**
Mean value (x):	2.6	4.2	7.7
Range of values:	2.3 - 3.1	3.2 - 4.8	7.0 - 8.8
Expected result:	positive	positive	positive
% positive:	100%	100%	100%
% negative:	0%	0%	0%

*3 lots x 2 runs ** n lots x 1 run

b. Linearity/assay reportable range: Not applicable.

High dose Hook effect C.

Potential for a high dose Hook effect is a phenomenon that is inherent with one step "sandwich" assay designs: Very high concentrations of antigen in the patient sample bind to all available sites saturating them - on both the antibody-solid phase and the antibody-labeled conjugate and thereby prevent the "sandwich" formation. Under these conditions, the measured level of analyte may be significantly lower than the actual level present in the sample. The two-step immunoassay design of the Anti-SS-B ELISA (IgG) eliminates the adverse contribution of binding proteins, endogenous interfering substances and general matrix effects due to the extra wash step.

18

Image /page/18/Picture/1 description: The image shows a pattern of four black circles evenly spaced between two horizontal black lines. The circles are solid black and appear to be the same size and shape. The lines are also solid black and appear to be parallel to each other. The circles are positioned in the center between the two lines. The pattern is simple and symmetrical.

d. Traceability, Stability, Expected values (controls, calibrators or methods):

A recognized standard or reference material for anti-nuclear antibodies is not available. Results of this assay are given in ratios. The reactivity of the Anti-SS-B ELISA was verified using the ANA reference panel of the CDC Centers for Disease Control and Prevention, Atlanta, USA.

• Limit of detection: e.
• Not applicable.
• f. Analytical specificity:

Cross-reactivity: The quality of the antigen coated on the plates ensures a high specificity of the ELISA. Cross reactivity was investigated using a panel of 30 sera serologically positive for antibodies against rib.P-P, nRNP/Sm, Sm. SS-A, Scl-70, Jo-1, centromeres, Rubella virus, Measles virus, Herpes simplex virus type 1 and Borrelia burgdorferi. All 30 sera were negative in the Anti-SS-B ELISA (IgG), so no cross reactivity is expected.

Interference: To investigate the influence from hemoglobin, triglycerides and bilirubin, 5 different sera at different anti-SS-B concentrations (18 - 123 RU/ml) were spiked with potential interfering substances and were incubated with the test system according to the package insert. Interferences from rheumatoid factor were investigated similarly by spiking with a RF positive serum. The recovery in relation to the unspiked sample without interferent was calculated. The individual recovery was within the range of 92 - 116 %. No significant interference was observed for concentrations of up to 1000 mg/dl for hemoglobin, 2000 mg/dl for triglyceride, 40 mg/dl for billrubin and 500 IU/ml for rheumatoid factor.

• Assay cut-off: টা
  Ratio 1.0

• 2. Comparison studies:
  • Method comparison with predicate device: a.

A comparison study was performed using 275 clinically characterized samples (57 SLE, 64 Sjögren's syndrome, 23 systemic sclerosis, 1 SSc/SS, 4 SLE/SS, 15 fibromyalgia, 26 myositis, 35 RA, 30 borreliosis and 20 healthy), obtained from different sources from Europe and North America. The panel consisted of 55 men and 220 women. Age ranged from 12 to 83 years with an average age of 48 vears. Anti-SS-B antibodies are expected in either Sjögren's syndrome or SLE. The other disease groups serve as control cohorts. The samples were tested with the EUROIMMUN Anti-SS-B ELISA (IgG) and with the Inova Quanta Lite SS-B ELISA as the predicate device. Of the 3 discrepant samples negative in the EUROIMMUN test, one was from a Siögren's syndrome patient and the other 2 were from controls. The discrepant sample positive in the EUROIMMUN test was from a Sjögren's syndrome patient.

| All samples

n = 275		Predicate ELISA
		positive	negative
EUROIMMUN
Anti-SS-B ELISA (IgG)	positive	82	1
	negative	3	189
Negative agreement		$189 / 190 = 99.5%$	95% C.I.: 97.1% - 100.0%
Positive agreement		$82 / 85 = 96.5%$	95% C.I.: 90.0% - 99.3%
Overall agreement		$271 / 275 = 98.5%$	95% C.I.: 96.3% - 99.6%

• b. Matrix comparison:
  The usability of plasma was investigated using sample pairs each of serum and corresponding plasma (EDTA, Li-heparin, Citrate). Passing-Bablok regression was calculated for the comparison of serum to plasma. The regression equation is near the ideal correlation (intercept 0; slope 1.0) indicating equivalence of concentrations between serum and the corresponding plasma matrices. Coefficients of determination were found to be above 0.99 and %recovery compared toserum was in the range of 78 to 116 % (serum = 100 %).

	EDTA plasma	Li-heparin plasma	Citrate plasma
n	16	16	16
Regression equation
(y = plasma, x = serum)	$y = -1.56 + 1.05 x$	$y = -0.22 + 1.06 x$	$y = 0.55 + 1.01$
95% C.I. of intercept	-5.46 - 0.19	-3.29 - 2.28	-1.56 - 5.62
95% C.I. of slope	1.04 - 1.09	1.01 - 1.09	0.97 - 1.05
Coefficient of determination R2	0.9972	0.9968	0.9951
Mean %recovery	100 %	105 %	101 %
Range of %recovery	78 - 116 %	98 - 116 %	81 - 110 %

19

Image /page/19/Picture/1 description: The image shows a repeating pattern of black shapes against a white background. The pattern consists of a horizontal line with rounded shapes above and below it. The rounded shapes are evenly spaced and appear to be identical.

• Clinical studies: ന്
  Clinical studies were performed in cooperation with different sites. In total 1026 clinically characterized samples (88 from SS patients, 404 from SLE patients and 534 from control groups) were investigated for anti-SS-B antibodies (IgG). With the EUROIMMUN Anti-SS-B ELISA (IgG) a prevalence of 39.8% (95% C.I.: 29.5 - 50.8%) was found in Siggren's syndrome and a prevalence of 13.9% (95% C.I.: 10.6 - 17.6%) was found in SLE with a specificity of 98.1% (95% C.I.: 96.6 – 99.1%). The results are shown in the table below. 95% C.I. are calculated by the exact method.

• Clinical sensitivity: a.

No.	Panel	n	Anti-SS-B ELISA (IgG)
			positive	%	95% C.I.
1	Sjögren's syndrome	88	35	39.8%	29.5 – 50.8%
2	Systemic lupus erythematosus	404	56	13.9%	10.6 – 17.6%

Clinical specificity: b.

No.	Panel	n	Anti-SS-B ELISA (IgG)
			negative	%	95% C.I.
3	Rheumatoid arthritis	164	163	99.4%	96.6 - 100.0%
4	Systemic sclerosis	81	77	95.1%	87.8 - 98.6%
5	Polymyositis/dermatomyositis	151	149	98.7%	95.3 - 99.8%
6	Mixed connective tissue diseases	45	42	93.3%	81.7 - 98.6%
7	Other autoimmune diseases*	63	63	100.0%	94.3 - 100.0%
8	Borreliosis	30	30	100.0%	88.4 - 100.0%
	Total	534	524	98.1%	96.6 - 99.1%

from the following groups: AIH (n = 8), Grave's disease (n = 12), Hashimoto (n = 11), celiac disease (n = 11), Diabetes Type I (n = 12)

• Other clinical supportive data (when a. and b. are not applicable): C. Not applicable.
• 4. Clinical cut-off:

See Assay cut-off.

• Expected values/Reference range: 5.
  The levels of anti-SS-B antibodies (IgG) were analyzed in a panel of 200 samples from apparently healthy blood donors (120 men and 80 women with an average age of 40 y; age range: 19 - 68 y). The results are shown in the table below.

n	200
Positives	0
Negatives	200
Prevalence	0.0%
	Ratio
Lowest value	0.0
Highest value	0.2
Mean value	0.0
Std deviation	0.02

N. Proposed Labeling:

The labeling is sufficient and it satisfies the requirements of 21 CFR Part 809.10.

். Conclusion:

The submitted information in this premarket notification is complete and supports a substantial equivalence decision.

Michael Locke
Signature

Dir. Regulatory Affairs Title

12 June 2013 Date

20

Image /page/20/Figure/1 description: The image shows a pattern of alternating black and white shapes. The black shapes are oval or circular, and they are arranged in a row. The white shapes are rectangular, and they are arranged in a row above and below the black shapes. The pattern is symmetrical and repetitive.

ATTACHMENT I

PREMARKET NOTIFICATION 510(K) SAFETY AND EFFECTIVENESS SUMMARY (as required by 21 CFR § 807.92)

• A. 510(k)Number: · K123261
• Purpose for Submission: B. New device
• C. Measurand:
  • Anti-Scl-70 autoantibodies
• D. Type of Test: Qualitative enzyme immunoassay
• E. Applicant:
  • EUROIMMUN US INC.
• F. Proprietary and Established Names: EUROIMMUN Anti-Scl-70 ELISA (IgG)
• G. Requlatory Information:
  • Regulation: 1.
    • 21 CFR 866.5110 Antinuclear antibody immunological test system
  • 2. Classification: Class II
  • 3. Product code: LLL
  • র . Panel: Immunology
• H. Intended Use:
  • 1. Intended use(s):

The EUROIMMUN Anti-Scl-70 ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against Scl-70 in human serum and plasma (EDTA, Li-heparin, Citrate), It is used as an aid in the diagnosis of progressive systemic sclerosis, in conjunction with other laboratory and clinical findings.

• 2. Indication(s) for use: Same as intended use.
• 3. Special conditions for the use statement(s): For prescription use only.
• 4. Special instrument requirements:

Microwell plate reader capable of measuring OD at 450nm for dual wavelength readings. Device Description:

• l.
  The EUROIMMUN Anti-Scl-70 ELISA (IgG) consists of a microwell ELISA plate coated with Scl-70 antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution.

J. Substantial Equivalence Information:

• Predicate device name (s): 1.
  • Inova Quanta Lite Scl-70 ELISA
• 2. Predicate 510(k) number(s): K924898

21

Image /page/21/Picture/1 description: The image shows a pattern of alternating black and white shapes. There are four black, oval-like shapes evenly spaced between two horizontal black lines. The white spaces between the black shapes create a repeating pattern.

ന് Comparison with predicate:

| Similarities

Item	New Device	Predicate Device
Intended use	Detection of IgG antibodies to Scl-70	Same
Technology	ELISA	Same
Assay platform	96-well microtiter plates	Same
Calibration	Relative units	Same
Antigen	Purified Scl-70 antigen	Same
Conjugate	Anti-human IgG labeled with horseradish peroxidase	Same
Substrate	TMB	Same
Reagent preparation	All reagents, calibrator and controls are ready to use,
except for the wash buffer.	Same
Procedure	Sample incubation with micro-well antigen coated plate,
followed by a wash step, incubation with an anti-human
IgG enzyme conjugate; wash step, incubation with
substrate; then the addition of a stop solution and
reading at 450nm.	Same
Differences
Item	New Device	Predicate Device
Assay format	Qualitative	"Semi-quantitative" (using low positive control)
Sample dilution	1:201	1:101
Calibrators and controls	1 calibrator
2 controls: 1 positive, 1 negative	3 controls: 1 high positive, 1 low positive (used for
calculation of results), 1 negative
Wash buffer	10x concentrate	40x concentrate
Stop solution	0.5 M sulphuric acid	0.344 M sulphuric acid
Sample types	Serum or plasma (EDTA, Li-heparin, Citrate)	Serum
Reported results	Ratio	Units

K. Standard/Guidance Document Referenced (if applicable):

Guidance for Industry and FDA Staff: Recommendations for Antibody (ANA) Test System Premarket (510(k)) Submissions (January 22, 2009)

20 Units

L. Test Principle:

Cut off level

Patient samples are diluted 1:201 in sample buffer. 100 ul of each diluted patient sample and pre-diluted controls and calibrators are added to the antigen coated microtiter wells and incubated for 30 minutes at room temperature. After incubation the microtiter well strips are washed with wash buffer to remove unbound antibodies and 100 ul of the anti-human IgG enzyme conjugate reagent is added to each microtiter well. After an additional 30-minutes incubation at room temperature, the microfiter wells are again washed 3 times with 300 µl of wash buffer to remove any unbound enzyme conjugate and 100 µl of the chromogen substrate is added. The strips are incubated for 15 minutes at room temperature and 100 ul stop solution is added. The microtiter plates are placed in an ELISA reader and read at a wavelength of 450 nm and a reference wavelength of between 620 nm and 650 nm within 30 minutes.

M. Performance Characteristics (where applicable):

Analytical performance: 1.

• a. Precision/Reproducibility:
  Ratio 1.0

The reproducibility of the test was investigated using sera with different concentrations. Intra-assay reproducibility is based on 20 determinations and inter-assay reproducibility on 40 determinations performed in 10 different runs on 5 days with 2 runs per day, each run performed according to the package insert. The following results were obtained:

22

Intra-assay reproducibility
	Anti-Scl-70 ELISA (IgG)
n = 20
	Sample 1	Sample 2	Sample 3	Sample 4	Sample 5	Sample 6
Mean value (x):	6.4	4.2	2.1	1.4	0.8	0.4
Range of values:	5.8 - 6.7	3.8 - 4.5	2.0 - 2.3	1.3 - 1.5	0.7 - 0.9	0.3 - 0.4
Expected result:	positive	positive	positive	positive	negative	negative
% positive:	100%	100%	100%	100%	0%	0%
% negative:	0%	0%	0%	0%	100%	100%

Inter-assay reproducibility

| n = 10 x 4 | Anti-Scl-70 ELISA (IgG)
Ratio | | | | | |
|------------------|----------------------------------|-----------|-----------|-----------|-----------|-----------|
| | Sample 1 | Sample 2 | Sample 3 | Sample 4 | Sample 5 | Sample 6 |
| Mean value (x): | 6.2 | 4.2 | 1.9 | 1.2 | 0.8 | 0.3 |
| Range of values: | 5.7 - 7.0 | 3.7 - 4.7 | 1.7 - 2.2 | 1.1 - 1.4 | 0.7 - 0.9 | 0.2 - 0.4 |
| Expected result: | positive | positive | positive | positive | negative | negative |
| % positive: | 100% | 100% | 100% | 100% | 0% | 0% |
| % negative: | 0% | 0% | 0% | 0% | 100% | 100% |

The lot to lot reproducibility was investigated during the validaton and quality control of the kit using different lots with QC samples distributed over the measurement range. The following results were obtained:

Lot to lot reproducibility

	Anti-Scl-70 ELISA (IgG)
	Ratio
	Sample 1	Sample 2	Sample 3	Sample 4	Sample 5	Sample 6
n	6*	6*	6*	6*	6*	6*
Mean value (x):	4.5	4.7	3.6	0.3	0.9	1.1
Range of values:	4.0 - 5.0	4.1 - 5.4	3.3 - 3.8	0.3-0.3	0.8 - 0.9	1.0 - 1.2
Expected result:	positive	positive	positive	negative	negative	positive
% positive:	100%	100%	100%	0%	0%	100%
% negative:	0%	0%	0%	100%	100%	0%

| | | Anti-Scl-70 ELISA (IgG)
Ratio | |
|------------------|-----------|----------------------------------|-----------|
| | Sample 7 | Sample 8 | Sample 9 |
| n | 11** | 10** | 11** |
| Mean value (x): | 3.0 | 4.8 | 6.2 |
| Range of values: | 2.5 - 3.7 | 4.3 - 5.7 | 5.3 - 6.8 |
| Expected result: | negative | positive | positive |
| % positive: | 0% | 100% | 100% |
| % negative: | 100% | 0% | 0% |

*3 lots x 2 runs ** n lots x 1 run

b. Linearity/assay reportable range: Not applicable.

High dose Hook effect C.

Potential for a high dose Hook effect is a phenomenon that is inherent with one step "sandwich" assay designs: Very high concentrations of antigen in the patient sample bind to all available sites saturating them - on both the antibody-solid phase and the antibody-labeled conjugate and thereby prevent the "sandwich" formation. Under these conditions, the measured level of analyte may be significantly lower than the actual level present in the sample. The two-step immunoassay design of the Anti-Scl-70 ELISA (IgG) eliminates the adverse contribution of binding proteins, endogenous interfering substances and general matrix effects due to the extra wash step.

23

Image /page/23/Picture/1 description: The image shows a black and white pattern of alternating dark and light vertical bars. The dark bars are wider in the middle and narrower at the top and bottom, creating a rounded shape. The light bars are uniform in width and separate the dark bars. The pattern is repeated several times across the image.

d. Traceability. Stability, Expected values (controls, calibrators or methods):

A recognized standard or reference material for anti-nuclear antibodies is not available. Results of this assay are given in ratios. The reactivity of the Anti-Scl-70 ELISA was verified using the ANA reference panel of the CDC Centers for Disease Control and Prevention, Atlanta, USA.

• Limit of detection: e.
• Not applicable.
• f. Analytical specificity:

Cross-reactivity: The quality of the antigen coated on the plates ensures a high specificity of the ELISA. Cross reactivity was investigated using a panel of 30 sera serologically positive for antibodies aqainst rib.P-P, nRNP/Sm, Sm, SS-A, SS-B, Jo-1, centromeres, Rubella virus, Measles virus, Herpes simplex virus type 1 and Borrelia burgdorferi. All 30 sera were negative in the Anti-Scl-70 ELISA (IgG), so no cross reactivity is expected.

Interference: To investigate the influence from hemoglobin, triglycerides and bilirubin, 5 different sera at different anti-Scl-70 concentrations (17 - 100 RU/ml) were spiked with potential interfering substances and were incubated with the test system according to the package insert. Interferences from rheumatoid factor were investigated similarly by spiking with a RF positive serum. The recovery in relation to the unspiked sample without interferent was calculated. The individual recovery was within the range of 86 - 109 %. No significant interference was observed for concentrations of up to 1000 mg/dl for hemoglobin, 2000 mg/dl for triglyceride, 40 mg/dl for bilirubin and 500 lU/ml for rheumatoid factor.

• Assay cut-off:
• Ratio 1.0

ರು

2. Comparison studies:

• Method comparison with predicate device: a.
  A comparison study was performed using 309 clinically characterized samples (158 systemic sclerosis, 51 Sjögren's syndrome, 15 fibromyalgia, 35 RA, 30 borreliosis and 20 healthy), obtained from different sources from Europe and North America. The panel consisted of 62 men and 247 women. Age ranged from 14 to 88 years with an average age of 53 years. Anti-Scl-70 antibodies are expected in systemic sclerosis. The other disease groups serve as control cohorts. The samples were tested with the EUROIMMUN Anti-Scl-70 ELISA (IgG) and with the Inova Quanta Lite Scl-70 ELISA as the predicate device.

| All samples

n = 309		Predicate ELISA
		positive	negative
EUROIMMUN
Anti-Scl-70 ELISA (IgG)	positive	125	0
EUROIMMUN
Anti-Scl-70 ELISA (IgG)	negative	0	184
Negative agreement	$184 / 184 = 100.0%$	95% C.I.: 98.0%	100.0%
Positive agreement	$125 / 125 = 100.0%$	95% C.I.: 97.1%	100.0%
Overall agreement	$309 / 309 = 100.0%$	95% C.I.: 98.8%	100.0%

• b. Matrix comparison:
  The usability of plasma was investigated using sample pairs each of serum and corresponding plasma (EDTA, Li-heparin, Citrate). Passing-Bablok regression was calculated for the comparison of serum to plasma. The regression equation is near the ideal correlation (intercept 0; slope 1.0) indicating equivalence of concentrations between serum and the corresponding plasma matrices. Coefficients of determination were found to be above 0.98 and %recovery compared toserum was in the range of 86 to 113 % (serum = 100 %).

	EDTA plasma	Li-heparin plasma	Citrate plasma
n	16	16	16
Regression equation
(y = plasma, x = serum)	$y = -0.58 + 1.00 x$	$y = 0.17 + 1.02 x$	$y = -0.87 + 0.97$
95% C.I. of intercept	-3.25 - 2.24	-1.99 - 1.64	-2.02 - 1.35
95% C.I. of slope	0.94 - 1.06	0.98 - 1.06	0.94 - 1.03
Coefficient of determination R2	0.9895	0.9930	0.9953
Mean %recovery	98 %	101 %	96 %
Range of %recovery	86 - 107 %	92 - 113 %	87 - 103 %

24

3. Clinical studies:

Clinical studies were performed in cooperation with different sites. In total 909 clinically characterized samples (280 from systemic sclerosis patients and 629 from control groups) were investigated for anti-Scl-70 antibodies (IgG). With the EUROMMUN Anti-Scl-70 ELISA (IgG) a prevalence of 23.2% (95% C.I.: 18.4 - 28.6%) was found in systemic sclerosis, a prevalence of 59.4% (95% C.I.: 48.9 - 69.3%) was found in diffuse systemic sclerosis and a prevalence of 5.3% (95% C.I.: 2.0 - 11.2%) was found in limited systemic sclerosis with a specificity of 99.8% (95% C.I.: 99.1 - 100.0%).The results are shown in the table below. 95% C.I. are calculated by the exact method.

• a. Clinical sensitivity:

No.	Panel	n	positive	%	95% C.I.
1	Systemic sclerosis	280	65	23.2%	18.4 – 28.6%
1a	Diffuse systemic sclerosis (from panel 1)	96	57	59.4%	48.9 – 69.3%
1b	Limited systemic sclerosis (from panel 1)	113	6	5.3%	2.0 – 11.2%

b. Clinical specificity:

No.	Panel	n	negative	%	95% C.I.
2	Systemic lupus erythematosus	213	213	100.0%	98.3 - 100.0%
3	Polymyositis/dermatomyositis	26	26	100.0%	86.8 - 100.0%
4	Rheumatoid arthritis	164	163	99.4%	96.6 - 100.0%
5	Sjögren's syndrome	88	88	100.0%	95.9 - 100.0%
6	Mixed connective tissue diseases	45	45	100.0%	92.1 - 100.0%
7	Other autoimmune diseases*	63	63	100.0%	94.3 - 100.0%
8	Borreliosis	30	30	100.0%	88.4 - 100.0%
	Total	629	628	99.8%	99.1 - 100.0%

ffrom the following groups: AIH (n = 8), Grave's disease (n = 12), Hashimoto (n = 11), celiac disease (n = 11), Diabetes Type I (n = 12)

Other clinical supportive data (when a. and b. are not applicable): C. Not applicable.

র্ব : Clinical cut-off:

See Assay cut-off.

5. Expected values/Reference range:

The levels of anti-Scl-70 antibodies (IgG) were analyzed in a panel of 200 samples from apparently healthy blood donors (120 men and 80 women with an average age of 40 y; age range: 19 - 68 y). The results are shown in the table below.

n	200
Positives	0
Negatives	200
Prevalence	0.0%
	Ratio
Lowest value	0.0
Highest value	0.1
Mean value	0.0
Std deviation	0.01

N. Proposed Labeling:

The labeling is sufficient and it satisfies the requirements of 21 CFR Part 809.10.

O. Conclusion:

The submitted information in this premarket notification is complete and supports a substantial equivalence decision.

Michael Locke
Signature

Dir. Requlatory Affairs

12 June 2013 Date

25

Image /page/25/Picture/1 description: The image shows a close-up of a series of dark, rounded shapes arranged in a row. The shapes are evenly spaced and appear to be three-dimensional. The background is light, creating a strong contrast with the dark shapes. The shapes are positioned between two dark horizontal lines.

ATTACHMENT 1

PREMARKET NOTIFICATION 210(K) SAFETY AND EFFECTIVENESS SUMMARY (as required by 21 CFR § 807.92)

• 510(k)Number: A. K123261
• Purpose for Submission: B. New device
• C. Measurand:
• Anti-Centromeres autoantibodies D. Type of Test:
  • Qualitative enzyme immunoassay
• E. Applicant:

.

H.

• EUROIMMUN US INC.
• F. Proprietary and Established Names: EUROIMMUN Anti-Centromeres ELISA (IgG)
• Regulatory Information: ઉ.
  • Regulation: 1.
    • 21 CFR 866.5110 Antinuclear antibody immunological test system
  • 2. Classification: Class II
  • 3. Product code:
  • LJM 4.
  • Panel:
  • Immunology Intended Use:
  • 1. Intended use(s):

The EUROIMMUN Anti-Centromeres ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against Centromeres in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of limited form of progressive systemic sclerosis (CREST syndrome), in conjunction with other laboratory and clinical findings.

• 2. Indication(s) for use: Same as intended use.
• 3. Special conditions for the use statement(s): For prescription use only.
• Special instrument requirements: 4.

Microwell plate reader capable of measuring OD at 450nm for dual wavelength readings.

• l. Device Description:
  The EUROIMMUN Anti-Centromeres ELISA (IgG) consists of a microwell ELISA plate coated with Centromeres antigen, calibrator, positive control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution.

• J. Substantial Equivalence Information:

  • Predicate device name (s): 1. Inova Quanta Lite Centromeres ELISA
  • 2. Predicate 510(k) number(s): KOO3AEa

26

Image /page/26/Picture/1 description: The image shows a pattern of alternating black and white shapes. There are four black oval shapes that are evenly spaced. The black ovals are sandwiched between two black horizontal lines.

3. Comparison with predicate:

Similarities
Item	New Device	Predicate Device
Intended use	Detection of IgG antibodies to Centromeres	Same
Technology	ELISA	Same
Assay platform	96-well microtiter plates	Same
Calibration	Relative units	Same
Conjugate	Anti-human IgG labeled with horseradish peroxidase	Same
Substrate	TMB	Same
Reagent preparation	All reagents, calibrator and controls are ready to use,
except for the wash buffer.	Same
Procedure	Sample incubation with micro-well antigen coated plate,
followed by a wash step, incubation with an anti-human
IgG enzyme conjugate; wash step, incubation with
substrate; then the addition of a stop solution and
reading at 450nm.	Same
Differences
Item	New Device	Predicate Device
Assay format	Qualitative	"Semi-quantitative" (using low positive control)
Antigen	1:201	Recombinant CENP-A and CENP-B
Sample dilution	1 calibrator	1:101
Calibrators and
controls	10x concentrate	3 controls: 1 high positive, 1 low positive (used for
calculation of results), 1 negative
Wash buffer	0.5 M sulphuric acid	40x concentrate
Stop solution	Serum or plasma (EDTA, Li-heparin, Citrate)	0.344 M sulphuric acid
Sample types	Ratio	Serum
Reported results	Ratio 1.0	Units
Cut off level	Qualitative	20 Units

K. Standard/Guidance Document Referenced (if applicable):

Guidance for Industry and FDA Staff: Recommendations for Antibody (ANA) Test System Premarket (510(k)) Submissions (January 22, 2009)

L. Test Principle:

Patient samples are diluted 1:201 in sample buffer. 100 ul of each diluted patient sample and pre-diluted controls and calibrators are added to the antigen coated microtiter wells and incubated for 30 minutes at room temperature. After incubation the microtiter well strips are washed with wash buffer to remove unbound antibodies and 100 µl of the anti-human IgG enzyme conjugate reagent is added to each microtiter well. After an additional 30-minutes incubation at room temperature, the microtiter wells are aqain washed 3 times with 300 ul of wash buffer to remove any unbound enzyme coniugate and 100 ul of the chromogen substrate is added. The strips are incubated for 15 minutes at room temperature and 100 ul stop solution is added. The microtiter plates are placed in an ELISA reader and read at a wavelength of 450 nm and a reference wavelength of between 620 nm and 650 nm within 30 minutes.

M. Performance Characteristics (where applicable):

• 1. Analytical performance:
  • a. Precision/Reproducibility:

The reproducibility of the test was investigated using sera with different concentrations. Intra-assay reproducibility is based on 20 determinations and inter-assay reproducibility on 40 determinations performed in 10 different runs on 5 days with 2 runs per day, each run performed according to the package insert. The following results were obtained:

27

Image /page/27/Picture/0 description: The image shows a black and white illustration of a ladder. The ladder has two long, horizontal rails that run parallel to each other. There are four rungs connecting the two rails. The rungs are evenly spaced and have a rounded shape.

| Intra-assay reproducibility | | Anti-Centromeres ELISA (IgG)
Ratio | | | | | |
|-----------------------------|-----------|---------------------------------------|-----------|-----------|-----------|-----------|----------|
| n = 20 | | Sample 1 | Sample 2 | Sample 3 | Sample 4 | Sample 5 | Sample 6 |
| Mean value (x): | 4.5 | 2.9 | 1.5 | 1.2 | 0.6 | 0.4 | |
| Range of values: | 4.4 - 4.6 | 2.7 - 2.9 | 1.5 - 1.6 | 1.1 - 1.2 | 0.6 - 0.6 | 0.3 - 0.4 | |
| Expected result:. | positive | positive | positive | positive | negative | negative | |
| % positive: | 100% | 100% | 100% | 100% | 0% | 0% | |
| % negative: | 0% | 0% | 0% | 0% | 100% | 100% | |

Inter-assay reproducibility

| n = 10 x 4 | Anti-Centromeres ELISA (IgG)
Ratio | | | | | |
|------------------|---------------------------------------|-----------|-----------|-----------|-----------|-----------|
| | Sample 1 | Sample 2 | Sample 3 | Sample 4 | Sample 5 | Sample 6 |
| Mean value (x): | 4.7 | 3.0 | 1.5 | 1.2 | 0.7 | 0.4 |
| Range of values: | 4.1 - 5.0 | 2.8 - 3.3 | 1.4 - 1.6 | 1.1 - 1.3 | 0.6 - 0.7 | 0.3 - 0.4 |
| Expected result: | positive | positive | positive | positive | negative | negative |
| % positive: | 100% | 100% | 100% | 100% | 0% | 0% |
| % negative: | 0% | 0% | 0% | 0% | 100% | 100% |

The lot to lot reproducibility was investigated during the validaton and quality control of the kit using different lots with QC samples distributed over the measurement range. The following results were obtained:

Lot to lot reproducibility

	Anti-Centromeres ELISA (IgG)
	Ratio
	Sample 1	Sample 2	Sample 3	Sample 4	Sample 5	Sample 6
n	6*	6*	6*	6*	6*	6*
Mean value (x):	4.1	4.9	3.9	0.3	0.9	1.1
Range of values:	3.8 - 4.5	4.6 - 5.3	3.7 - 4.2	0.3 - 0.3	0.8 - 0.9	1.1 - 1.2
Expected result:	positive	positive	positive	negative	negative	positive
% positive:	100%	100%	100%	0%	0%	100%
% negative:	0%	0%	0%	100%	100%	0%

| | Anti-Centromeres ELISA (IgG)
Ratio | | | |
|------------------|---------------------------------------|-----------|-----------|-----------|
| | Sample 7 | Sample 8 | Sample 9 | Sample 10 |
| n | 8** | 10** | 11** | 10** |
| Mean value (x): | 2.8 | 6.2 | 6.8 | 8.4 |
| Range of values: | 2.2 - 3.3 | 5.5 - 7.0 | 4.9 - 8.3 | 7.8 - 9.1 |
| Expected result: | positive | positive | positive | positive |
| % positive: | 100% | 100% | 100% | 100% |
| % negative: | 0% | 0% | 0% | 0% |

*3 lots x 2 runs ** n lots x 1 run

• Linearity/assay reportable range: b. Not applicable.
  High dose Hook effect C.

Potential for a high dose Hook effect is a phenomenon that is inherent with one step "sandwich" assay designs: Very high concentrations of antigen in the patient sample bind to all available sites saturating them - on both the antibody-solid phase and the antibody-labeled conjugate and thereby prevent the "sandwich" formation. Under these conditions, the measured level of analyte may be significantly lower than the actual level present in the sample. The two-step immunoassay design of the Anti-Centromeres ELISA (IgG) eliminates the adverse contribution of binding proteins. endogenous interfering substances and general matrix effects due to the extra wash step.

28

d. Traceability, Stability, Expected values (controls, calibrators or methods):

A recognized standard or reference material for anti-nuclear antibodies is not available. Results of this assay are given in ratios. The reactivity of the Anti-Centromeres ELISA was verified using the ANA reference panel of the CDC Centers for Disease Control and Prevention, Atlanta, USA.

• Limit of detection: રે. Not applicable.
• f. Analytical specificity:

Cross-reactivity: The quality of the antigen coated on the plates ensures a high specificity of the ELISA. Cross reactivity was investigated using a panel of 30 sera serologically positive for antibodies against rib.P-P, nRNP/Sm, Sm, SS-A, Scl-70, Jo-1, Rubella virus, Measles virus, Herpes simplex virus type 1 and Borrelia burgdorferi. All 30 sera were negative in the Anti-Centromeres ELISA (IgG), so no cross reactivity is expected.

Interference: To investigate the influence from hemoglobin, triglycerides and billrubin, 5 different sera at different anti-Centromeres concentrations (12 - 106 RU/ml) were spiked with potential interfering substances and were incubated with the test system according to the package insert. Interferences from rheumatoid factor were investigated similarly by spiking with a RF positive serum. The recovery in relation to the unspiked sample without interferent was calculated. The individual recovery was within the range of 91 - 111 %. No significant interference was observed for concentrations of up to 1000 mg/dl for hemoglobin, 2000 mg/dl for triglyceride, 40 mg/dl for billrubin and 500 IU/ml for rheumatoid factor.

• ರು Assay cut-off:
  Ratio 1.0

Comparison studies:

• Method comparison with predicate device: a.
  A comparison study was performed using 297 clinically characterized samples (144 systemic sclerosis, 2 CREST, 51 Sjögren's syndrome, 15 fibromyalgia, 35 RA, 30 borreliosis and 20 healthy), obtained from different sources from Europe and North America. The panel consisted of 61 men and 230 women. Age ranged from 2 to 87 vears with an average age of 50 years. Anti-centromeres antibodies are expected in systemic sclerosis. The other disease groups serve as control cohorts. The samples were tested with the EUROIMMUN Anti-Centromeres ELISA (IgG) and with the Inova Quanta Lite Centromere ELISA as the predicate device. All of the 5 discrepant samples were from control groups.

| All samples

n = 297		Predicate ELISA
		positive	negative
EUROIMMUN
Anti-Centromeres ELISA (IgG)	positive	73	0
	negative	5	219
Negative agreement	$219 / 219 = 100.0%$	95% C.I.: 98.3%	- 100.0%
Positive agreement	$73 / 78 = 93.6%$	95% C.I.: 85.7%	- 97.9%
Overall agreement	$292 / 297 = 98.3%$	95% C.I.: 96.1%	- 99.5%

b. Matrix comparison:

The usability of plasma was investigated using sample pairs each of serum and corresponding plasma (EDTA, Li-heparin, Citrate). Passing-Bablok regression was calculated for the comparison of serum to plasma. The regression equation is near the ideal correlation (intercept 0; slope 1.0) indicating equivalence of concentrations between serum and the corresponding plasma matrices. Coefficients of determination were found to be above 0.99 and %recovery compared toserum was in the range of 84 to 108 % (serum = 100 %).

	EDTA plasma	Li-heparin plasma	Citrate plasma
n	16	16	16
Regression equation
(y = plasma, x = serum)	$y = -1.54 + 1.02 x$	$y = 0.02 + 0.92 x$	$y = -0.99 + 0.98$
95% C.I. of intercept	-3.85 – -0.06	-2.13 – 0.99	-2.48 – 0.69
95% C.I. of slope	0.98 – 1.08	0.89 – 0.96	0.94 – 1.01
Coefficient of determination R2	0.9958	0.9973	0.9978
Mean %recovery	98 %	93 %	96 %
Range of %recovery	84 – 108 %	83 – 101 %	88 – 107 %

29

3. Clinical studies:

Clinical studies were performed in cooperation with different sites. In total 877 clinically characterized samples (280 from systemic sclerosis patients and 597 from control groups) were investigated for anticentromeres antibodies (IgG). With the EUROIMMUN Anti-Centromeres ELISA (IgG) a prevalence of 19.0% (95% C.I.: 13.9 - 24.9%) was found in systemic sclerosis, a prevalence of 7.3% (95% C.I.: 3.0 -14.4%) was found in diffuse systemic sclerosis and a prevalence of 74.3% (95% C.I.: 65.3 - 82.1%) was found in limited systemic sclerosis with a specificity of 99.0% (95% C.I.; 97.8 - 99.6%). The results are shown in the table below. 95% C.I. are calculated by the exact method.

• Clinical sensitivity: a.

No.	Panel	n	Anti-Centromeres ELISA (IgG)
			positive	%	95% C.I.
1	Systemic sclerosis	280	105	37.5%	31.8 - 43.5%
1a	Diffuse systemic sclerosis (from panel 1)	96	7	7.3%	3.0 - 14.4%
1b	Limited systemic sclerosis (from panel 1)	113	84	74.3%	65.3 - 82.1%

b. Clinical specificity:

No.	Panel	n.	negative	%	95% C.I.
2	Systemic lupus erythematosus	181	180	99.4%	97.0 - 100.0%
3	Polymyositis/dermatomyositis	26	26	100.0%	86.8 - 100.0%
4	Rheumatoid arthritis	164	163	99.4%	96.6 - 100.0%
5	Sjögren's syndrome	88	85	96.6%	90.4 - 99.3%
6	Mixed connective tissue diseases	45	44	97.8%	88.2 - 99.9%
7	Other autoimmune diseases*	63	63	100.0%	94.3 - 100.0%
8	Borreliosis	30	30	100.0%	88.4 - 100.0%
	Total	597	591	99.0%	97.8 - 99.6%

*from the following groups: AIH (n = 8), Grave's disease (n = 12), Hashimoto (n = 11), celiac disease (n = 11), Diabetes Type I (n = 12)

• Other clinical supportive data (when a. and b. are not applicable): C.
  • Not applicable.
• 4. Clinical cut-off:

See Assay cut-off.

• ട്. Expected values/Reference range:
  The levels of anti-centromeres antibodies (IgG) were analyzed in a panel of 200 samples from apparently healthy blood donors (120 men and 80 women with an average age of 40 y; age range: 19 - 68 y). The results are shown in the table below.

n	200
Positives	1
Negatives	199
Prevalence	0.5%
	Ratio
Lowest value	0.0
Highest value	3.0
Mean value	0.1
Std deviation	0.21

· N. Proposed Labeling:

The labeling is sufficient and it satisfies the requirements of 21 CFR Part 809.10.

O. Conclusion:

The submitted information in this premarket notification is complete and supports a substantial equivalence decision.

Michael Locke
Signature

Dir. Regulatory Affairs

12 June 2013 Dote

30

Image /page/30/Figure/1 description: The image shows a pattern of alternating black and white shapes. The black shapes appear to be horizontal bars running across the top and bottom of the image. In between the bars are a series of black oval shapes, separated by white space. There are four black oval shapes in total.

ATTACHMENT I

PREMARKET NOTIFICATION 210(K) SAFETY AND EFFECTIVENESS SUMMARY (as required by 21 CFR § 807.92)

• A. 510(k)Number:

K123261

• Purpose for Submission: B. New device
• C. Measurand:
  • Anti-Jo-1 autoantibodies
• D. Type of Test: Qualitative enzyme immunoassay
• E. Applicant:
  • EUROIMMUN US INC.
• F. Proprietary and Established Names: EUROIMMUN Anti-Jo-1 ELISA (IgG)

ઉં. Requlatory Information:

• 1. Regulation:
  • 21 CFR 866.5110 Antinuclear antibody immunological test system
• 2. Classification: Class II
• 3. Product code: LLL
• ধ . Panel:

H.

• Immunology Intended Use:
• Intended use(s): 1.

The EUROIMMUN Anti-Jo-1 ELISA (IqG) test kit is intended for the qualitative determination of IgG class autoantibodies against Jo-1 in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of polymyositis and dermatomyositis, in conjunction with other laboratory and clinical findings.

• 2. Indication(s) for use: Same as intended use.
• 3. Special conditions for the use statement(s): For prescription use only.

4. Special instrument requirements:

Microwell plate reader capable of measuring OD at 450nm for dual wavelength readings.

l. Device Description:

The EUROIMMUN Anti-Jo-1 ELISA (IgG) consists of a microwell ELISA plate coated with Jo-1 antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution and stop solution.

۔ ل۔ Substantial Equivalence Information:

• Predicate device name (s): 1.
  • Inova Quanta Lite Jo-1 ELISA
• 2. Predicate 510(k) number(s): Kazesez

31

Image /page/31/Picture/1 description: The image shows a pattern of alternating black and white shapes. There are four black oval shapes that are separated by white rectangular shapes. The black shapes are connected by two horizontal black lines, one at the top and one at the bottom. The pattern is symmetrical and evenly spaced.

3. Comparison with predicate:

Similarities
Item	New Device	Predicate Device
Intended use	Detection of IgG antibodies to Jo-1	Same
Technology	ELISA	Same
Assay platform	96-well microtiter plates	Same
Calibration	Relative units	Same
Antigen	Purified Jo-1 antigen	Same
Conjugate	Anti-human IgG labeled with horseradish peroxidase	Same
Substrate	TMB	Same
Reagent preparation	All reagents, calibrator and controls are ready to use,
except for the wash buffer.	Same
Procedure	Sample incubation with micro-well antigen coated plate,
followed by a wash step, incubation with an anti-human
IgG enzyme conjugate; wash step, incubation with
substrate; then the addition of a stop solution and
reading at 450nm.	Same
Differences
Item	New Device	Predicate Device
Assay format	Qualitative	"Semi-quantitative" (using low positive control)
Sample dilution	1:201	1:101
Calibrators and	1 calibrator	3 controls: 1 high positive, 1 low positive (used for
controls	2 controls: 1 positive, 1 negative	calculation of results), 1 negative
Wash buffer	10x concentrate	40x concentrate
Stop solution	0.5 M sulphuric acid	0.344 M sulphuric acid
Sample types	Serum or plasma (EDTA, Li-heparin, Citrate)	Serum
Reported results	Ratio	Units
Cut off level	Ratio 1.0	20 Units

K. Standard/Guidance Document Referenced (if applicable):

Guidance for Industry and FDA Staff: Recommendations for Antibody (ANA) Test System Premarket (510(k)) Submissions (January 22, 2009)

L. Test Principle:

Patient samples are diluted 1:201 in sample buffer, 100 µl of each diluted patient sample and pre-diluted controls and calibrators are added to the antigen coated microtiter wells and incubated for 30 minutes at room temperature. After incubation the microtiter well strips are washed with wash buffer to remove unbound antibodies and 100 ul of the anti-human IgG enzyme coniugate reagent is added to each microtiter well. After an additional 30-minutes incubation at room temperature, the microtiter wells are again washed 3 times with 300 ul of wash buffer to remove any unbound-enzyme conjugate and 100 µl of the chromogen substrate is added. The strips are incubated for 15 minutes at room temperature and 100 ul stop solution is added. The microtiter plates are placed in an ELISA reader and read at a wavelength of 450 nm and a reference wavelength of between 620 nm and 650 nm within 30 minutes.

M. Performance Characteristics (where applicable):

Analytical performance: 1.

• Precision/Reproducibility: a.
  The reproducibility of the test was investigated using sera with different concentrations. Intra-assay reproducibility is based on 20 determinations and inter-assay reproducibility on 40 determinations performed in 10 different runs on 5 days with 2 runs per day, each run performed according to the package insert. The following results were obtained:

32

Image /page/32/Picture/0 description: The image shows a black and white pattern. The pattern consists of four black shapes that are separated by white spaces. The black shapes are connected by two horizontal black lines, one at the top and one at the bottom. The shapes are symmetrical and have a curved appearance.

Intra-assay reproducibility

| | Anti-Jo-1 ELISA ELISA (IgG)
Ratio | | | | | |
|------------------|--------------------------------------|-----------|-----------|-----------|-----------|-----------|
| n = 20 | | | | | | |
| | Sample 1 | Sample 2 | Sample 3 | Sample 4 | Sample 5 | Sample 6 |
| Mean value (x): | 6.4 | 4.3 | 1.8 | 1.0 | 0.8 | 0.4 |
| Range of values: | 6.1 - 6.8 | 4.1 - 4.5 | 1.6 - 1.9 | 1.0 - 1.1 | 0.7 - 0.8 | 0.3 - 0.4 |
| Expected result: | positive | positive | positive | positive | negative | negative |
| % positive: | 100% | 100% | 100% | 100% | 0% | 0% |
| % negative: | 0% | 0% | 0% | 0% | 100% | 100% |

Inter-assay reproducibility

| n = 10 x 4 | Anti-Jo-1 ELISA (IgG)
Ratio | | | | | |
|------------------|--------------------------------|-----------|-----------|-----------|-----------|-----------|
| | Sample 1 | Sample 2 | Sample 3 | Sample 4 | Sample 5 | Sample 6 |
| Mean value (x): | 6.0 | 4.2 | 2.2 | 1.1 | 0.8 | 0.4 |
| Range of values: | 5.6 - 6.6 | 3.7 - 4.6 | 2.0 - 2.3 | 1.0 - 1.2 | 0.7 – 0.8 | 0.3 - 0.4 |
| Expected result: | positive | positive | positive | positive | negative | negative |
| % positive: | 100% | 100% | 100% | 100% | 0% | 0% |
| % negative: | 0% | 0% | 0% | 0% | 100% | 100% |

The lot to lot reproducibility was investigated during the validaton and quality control of the kit using different lots with QC samples distributed over the measurement range. The following results were obtained:

Lot to lot reproducibility

| | Anti-Jo-1 ELISA (IgG) Ratio | | | | | | | | Anti-Jo-1 ELISA (IgG)
Ratio | | | |
|------------------|-----------------------------|-----------|-----------|-----------|-----------|-----------|------------------|-----------|--------------------------------|-----------|-----------|--|
| | Sample 1 | Sample 2 | Sample 3 | Sample 4 | Sample 5 | Sample 6 | | Sample 7 | Sample 8 | Sample 9 | Sample 10 | |
| n | 6* | 6* | 6* | 6* | 6* | 6* | n | 10** | 11** | 11** | 11** | |
| Mean value (x): | 5.1 | 3.9 | 2.0 | 0.3 | 0.9 | 1.1 | Mean value (x): | 2.6 | 4.0 | 7.4 | 8.2 | |
| Range of values: | 4.6 - 5.4 | 3.5 - 4.4 | 1.9 - 2.2 | 0.3 - 0.4 | 0.9 - 0.9 | 1.1 - 1.2 | Range of values: | 1.9 - 3.1 | 3.2 – 4.6 | 6.4 - 8.6 | 7.7 - 8.9 | |
| Expected result: | positive | positive | positive | negative | negative | positive | Expected result: | positive | positive | positive | positive | |
| % positive: | 100% | 100% | 100% | 0% | 0% | 100% | % positive: | 100% | 100% | 100% | 100% | |
| % negative: | 0% | 0% | 0% | 100% | 100% | 0% | % negative: | 0% | 0% | 0% | 0% | |

*3 lots x 2 runs ** n lots x 1 run

• Linearity/assay reportable range: b. Not applicable.
• High dose Hook effect C.

Potential for a high dose Hook effect is a phenomenon that is inherent with one step "sandwich" assay designs: Very high concentrations of antigen in the patient sample bind to all available sites saturating them - on both the antibody-solid phase and the antibody-labeled conjugate and thereby prevent the "sandwich" formation. Under these conditions, the measured level of analyte may be significantly lower than the actual level present in the sample. The two-step immunoassay design of the Anti-Jo-1 ELISA (IgG) eliminates the adverse contribution of binding proteins, endogenous interfering substances and general matrix effects due to the extra wash step.

33

Image /page/33/Picture/1 description: The image shows a black and white illustration of a decorative border. The border consists of a series of rounded shapes, resembling beads or small bulbs, positioned between two horizontal lines. The shapes are evenly spaced and create a repeating pattern along the length of the border.

d. Traceability, Stability, Expected values (controls, calibrators or methods):

A recognized standard or reference material for anti-nuclear antibodies is not available. Results of this assay are given in ratios. The reactivity of the Anti-Jo-1 ELISA was verified using the ANA reference panel of the CDC Centers for Disease Control and Prevention, Atlanta, USA.

• Limit of detection: e.
• Not applicable. f. Analytical specificity:

Cross-reactivity: The quality of the antigen coated on the plates ensures a high specificity of the ELISA. Cross reactivity was investigated using a panel of 30 sera serologically positive for antibodies against rib.P-P, nRNP/Sm, Sm, SS-A, SS-B, Scl-70, centromeres, Rubella virus, Measles virus, Herpes simplex virus type 1 and Borrelia burgdorferi. All 30 sera were negative in the Anti-Jo-1 ELISA (IgG), so no cross reactivity is expected.

Interference: To investigate the influence from hemoglobin, triglycerides and bilirubin, 5 different sera at different anti-Jo-1 concentrations (15 - 106 RU/ml) were spiked with potential interfering substances and were incubated with the test system according to the package insert. Interferences from rheumatoid factor were investigated similarly by spiking with a RF positive serum. The recovery in relation to the unspiked sample without interferent was calculated. The individual recovery was within the range of 91 - 122 %. No significant interference was observed for concentrations of up to 1000 mg/dl for hemoglobin, 2000 mg/dl for triglyceride, 40 mg/dl for bilirubin and 500 lU/ml for rheumatoid factor.

• Assay cut-off: ರು
• Ratio 1.0

2. Comparison studies:

• Method comparison with predicate device: a.
  A comparison study was performed using 297 clinically characterized samples (143 myositis, 3 PM/SSc, 51 Sjögren's syndrome, 15 fibromyalgia, 35 RA, 30 borreliosis and 20 healthy), obtained from different sources from Europe and North America. The panel consisted of 75 men and 221 women. Age ranged from 14 to 84 years with an average age of 52 years. Anti-Jo-1 antibodies are expected in myositis. The other disease groups serve as control cohorts. The samples were tested with the EUROIMMUN Anti-Jo-1 ELISA (IgG) and with the Inova Quanta Lite Jo-1 ELISA as the predicate device. Both discrepant samples were from myositis patients.

| All samples

n = 297		Predicate ELISA
		positive	negative
EUROIMMUN
Anti-Jo-1 ELISA (IgG)	positive	64	1
	negative	1	231
Negative agreement		231 / 232 = 99.6%	95% C.I.: 97.6% - 100.0%
Positive agreement		64 / 65 = 98.5%	95% C.I.: 91.7% - 100.0%
Overall agreement		295 / 297 = 99.3%	95% C.I.: 97.6% - 99.9%

• b. Matrix comparison:
  The usability of plasma was investigated using sample pairs each of serum and corresponding plasma (EDTA, Li-heparin, Citrate). Passing-Bablok regression was calculated for the comparison of serum to plasma. The regression equation is near the ideal correlation (intercept 0; slope 1.0) indicating equivalence of concentrations between serum and the corresponding plasma matrices. Coefficients of determination were found to be above 0.98 and %recovery compared toserum was in the range of 85 to 117 % (serum = 100 %).

	EDTA plasma	Li-heparin plasma	Citrate plasma
n	15	15	15
Concentration range (serum)	ratio 0.3 - 9.2	ratio 0.3 - 9.2	ratio 0.3 - 9.2
Concentration range (plasma)	ratio 0.3 - 8.7	ratio 0.4 - 8.5	ratio 0.3 - 8.9
Regression equation
(y = plasma, x = serum)	y = -0.09 + 0.99 x	y = 0.09 + 0.93 x	y = 0.73 + 0.99
95% C.I. of intercept	-3.77 - 1.83	-0.69 - 1.94	-0.49 - 2.66
95% C.I. of slope	0.93 - 1.10	0.89 - 0.99	0.96 - 1.01
Coefficient of determination R2	0.9856	0.9956	0.9982
Mean %recovery	101 %	95 %	103 %
Range of %recovery	85 - 117 %	87 - 113 %	94 - 114 %

34

3. Clinical studies:

Clinical studies were performed in cooperation with different sites. In total 876 clinically characterized samples (177 from myositis patients and 699 from control groups) were investigated for anti-Jo-1 antibodies (IgG). With the EUROIMMUN Anti-Jo-1 ELISA (IgG) a prevalence of 18.6% (95% C.I.: 13.2 – 25.2%) was found in myositis with a specificity of 99.6% (95% C.I.: 98.8 – 99.9%). The results are shown in the table below. 95% C.I. are calculated by the exact method. "

• a. Clinical sensitivity:

| NO. | Panel | | FLISA (IaG)
Anti-Jo-1 | | | |
|-----|----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|---------|--------------------------|--------|---------------------|--|
| | | | positive | | 95% C.I. | |
| | ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Polymvositis/dermatomyositis | 1 79 97 | | 18.6% | 25.2%
1 - 25.6 . | |
| No. | Panel | n | Anti-Jo-1 ELISA (IgG) | | | |
| | | | negative | % | 95% C.I. | |
| 2 | Systemic lupus erythematosus | 213 | 211 | 99.1% | 96.6 - 99.9% | |
| 3 | Rheumatoid arthritis | 164 | 163 | 99.4% | 96.6 - 100.0% | |
| 4 | Systemic sclerosis | 81 | 81 | 100.0% | 95.5 - 100.0% | |
| 5 | Sjögren's syndrome | 88 | 88 | 100.0% | 95.9 - 100.0% | |
| 6 | Mixed connective tissue diseases | 45 | 45 | 100.0% | 92.1 - 100.0% | |
| 7 | Fibromyalgia | 15 | 15 | 100.0% | 78.2 - 100.0% | |
| 8 | Other autoimmune diseases | 63 | 63 | 100.0% | 94.3 - 100.0% | |
| 9 | Borreliosis | 30 | 30 | 100.0% | 88.4 - 100.0% | |
| | Total | 699 | 696 | 99.6% | 98.8 - 99.9% | |

b. Clinical specificity:

Other clinical supportive data (when a. and b. are not applicable): C.

• Not applicable.
  ব : Clinical cut-off:

See Assay cut-off.

• 5. Expected values/Reference range:
     The levels of antibodies (IgG) were analyzed in a panel of 200 samples from apparently healthy blood donors (120 men and 80 women with an average age of 40 y; age range: 19 - 68 y). The results are shown in the table below.

n	200
Positives	0
Negatives	200
Prevalence	0.0%
	Ratio
Lowest value	0.0
Highest value	0.2
Mean value	0.1
Std deviation	0.03

N. Proposed Labeling:

The labeling is sufficient and it satisfies the requirements of 21 CFR Part 809.10.

O. Conclusion:

The submitted information in this premarket notification is complete and supports a substantial equivalence decision.

Michael Locke
Signature

Dir. Regulatory Affairs Title

12 June 2013 Date

35

Image /page/35/Figure/1 description: The image shows a black and white pattern. The pattern consists of a series of black shapes that are connected by thin black lines. The shapes are arranged in a row, and they are all the same size and shape. The shapes are symmetrical, with a rounded top and bottom and a narrow waist.

ATTACHMENT I

PREMARKET NOTIFICATION 200(K) SAFETY AND EFFECTIVENESS SUMMARY (as required by 21 CFR § 807.92)

• A. 510(k)Number:
  • K123261
• B. Purpose for Submission: · New device
• C. Measurand:
  • Anti-ribosomal P-proteins autoantibodies
• D. Type of Test:
  • Qualitative enzyme immunoassay
• E. Applicant: EUROIMMUN US INC.
• F. Proprietary and Established Names: EUROIMMUN Anti-ribosomal P-proteins ELISA (IgG)

G. Requlatory Information:

• 1. Regulation:
  • 21 CFR 866.5110 Antinuclear antibody immunological test system
• 2. Classification: Class II
• 3. Product code: MQA -
• র . Panel:

H.

l.

• Immunology
• Intended Use:
• 1. Intended use(s):

The EUROIMMUN Anti-ribosomal P-proteins ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against ribosomal P-proteins in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of systemic lupus erythematosus, in conjunction with other laboratory and clinical findings.

• 2. Indication(s) for use: Same as intended use.
• 3. Special conditions for the use statement(s): For prescription use only.

র্ব . Special instrument requirements:

Microwell plate reader capable of measuring OD at 450nm for dual wavelength readings. Device Description:

The EUROIMMUN Anti-ribosomal P-proteins ELISA (IgG) consists of a microwell ELISA plate coated with ribosomal P-proteins antigen, calibrator, positive and negative control, peroxidase-labelled anti-human IgG conjugate, sample buffer, wash buffer concentrate, TMB chromogen/substrate solution.

J. Substantial Equivalence Information:

• Predicate device name (s): 1.
  • Inova Quanta Lite Ribosomal P ELISA
• 2. Predicate 510(k) number(s): K981237

36

Image /page/36/Picture/1 description: The image shows the number 3 followed by the words "Comparison with predicate:". The text is black and the background is white. The text is left-aligned.

| Similarities

Item	New Device	Predicate Device
Intended use	Detection of IgG antibodies to ribosomal P-proteins	Same
Technology	ELISA	Same
Assay platform	96-well microtiter plates	Same
Calibration	Relative units	Same
Conjugate	Anti-human IgG labeled with horseradish peroxidase	Same
Substrate	TMB	Same
Reagent preparation	All reagents, calibrator and controls are ready to use,
except for the wash buffer.	Same
Procedure	Sample incubation with micro-well antigen coated plate,
followed by a wash step, incubation with an anti-human
IgG enzyme conjugate; wash step, incubation with
substrate; then the addition of a stop solution and
reading at 450nm.	Same
Differences
Item	New Device	Predicate Device
Assay format	Qualitative	"Semi-quantitative" (using low positive control)
Antigen	Purified ribosomal P antigen	Synthetic ribosomal P antigen
Sample dilution	1:201	1:101
Calibrators and
controls	1 calibrator
2 controls: 1 positive, 1 negative	3 controls: 1 high positive, 1 low positive (used for
calculation of results), 1 negative
Wash buffer	10x concentrate	40x concentrate
Stop solution	0.5 M sulphuric acid	0.344 M sulphuric acid
Sample types	Serum or plasma (EDTA, Li-heparin, Citrate)	Serum
Reported results	Ratio	Units
Cut off level	Ratio 1.0	20 Units

K. Standard/Guidance Document Referenced (if applicable):

Guidance for Industry and FDA Staff: Recommendations for Antibody (ANA) Test System Premarket (510(k)) Submissions (January 22, 2009)

L. Test Principle:

Patient samples are diluted 1:201 in sample buffer, 100 pl of each diluted patient sample and pre-diluted controls and calibrators are added to the antigen coated microtiter wells and incubated for 30 minutes at room temperature. After incubation the microtier well strips are washed with wash buffer to remove unbound antibodies and 100 ul of the anti-human IgG enzyme conjugate reagent is added to each microtiter well. After an additional 30-minutes incubation at room temperature, the microtiter wells are again washed 3 times with 300 ul of wash buffer to remove any unbound enzyme conjuqate and 100 ul of the chromogen substrate is added. The strips are incubated for 15 minutes at room temperature and 100 ul stop solution is added. The microtier placed in an ELISA reader and read at a wavelength of 450 nm and a reference wavelength of between 620 nm and 650 nm within 30 minutes.

Performance Characteristics (where applicable): M.

• Analytical performance: 1.
  • Precision/Reproducibility: a.

The reproducibility of the test was investigated using sera with different concentrations. Intra-assay reproducibility is based on 20 determinations and inter-assay reproducibility on 40 determinations performed in 10 different runs on 5 days with 2 runs per day, each run performed according to the package insert. The following results were obtained:

37

Image /page/37/Picture/0 description: The image shows a black and white illustration of a decorative border. The border consists of a series of oval or rounded shapes, evenly spaced and connected by horizontal lines above and below. The shapes are solid black, creating a stark contrast against the white background.

Intra-assay reproducibility

| | Anti-ribosomal P-proteins ELISA (IgG)
Ratio | | | | | |
|------------------|------------------------------------------------|-----------|-----------|-----------|-----------|-----------|
| n = 20 | | | | | | |
| | Sample 1 | Sample 2 | Sample 3 | Sample 4 | Sample 5 | Sample 6 |
| Mean value (x): | 5.5 | 3.6 | 1.9 | 1.0 | 0.6 | 0.4 |
| Range of values: | 5.1 - 6.0 | 3.2 - 4.1 | 1.8 - 2.1 | 1.0 - 1.1 | 0.6 - 0.7 | 0.3 - 0.5 |
| Expected result: | positive | positive | positive | positive | negative | negative |
| % positive: | 100% | 100% | 100% | 100% | 0% | 0% |
| % negative: | 0% | 0% | 0% | 0% | 100% | 100% |

Inter-assay reproducibility

| n = 10 x 4 | Anti-ribosomal P-proteins ELISA (IgG)
Ratio | | | | | |
|------------------|------------------------------------------------|-----------|-----------|-----------|-----------|-----------|
| | Sample 1 | Sample 2 | Sample 3 | Sample 4 | Sample 5 | Sample 6 |
| Mean value (x): | 5.7 | 3.8 | 1.9 | 1.1 | 0.7 | 0.4 |
| Range of values: | 5.3 - 6.1 | 3.5 - 4.1 | 1.7 - 2.2 | 1.0 - 1.3 | 0.6 - 1.0 | 0.3 - 0.4 |
| Expected result: | positive | positive | positive | positive | negative | negative |
| % positive: | 100% | 100% | 100% | 100% | 2.5% | 0% |
| % negative: | 0% | 0% | 0% | 0% | 97.5% | 100% |

The lot to lot reproducibility was investigated during the validaton and quality control of the kit using different lots with QC samples distributed over the measurement range. The following results were obtained:

Lot to lot reproducibility

| | Anti-ribosomal P-proteins ELISA (IgG)
Ratio | | | | | |
|------------------|------------------------------------------------|-----------|-----------|----------|-----------|-----------|
| | Sample 1 | Sample 2 | Sample 3 | Sample 4 | Sample 5 | Sample 6 |
| n | 6* | 6* | 6* | 6* | 6* | 6* |
| Mean value (x): | 3.2 | 4.5 | 2.2 | 0.3 | 0.9 | 1.1 |
| Range of values: | 2.8 – 3.6 | 4.2 - 4.8 | 2.0 - 2.4 | 0.3-0.4 | 0.8 - 0.9 | 1.0 - 1.2 |
| Expected result: | positive | positive | positive | negative | negative | positive |
| % positive: | 100% | 100% | 100% | 0% | 0% | 100% |
| % negative: | 0% | 0% | 0% | 100% | 100% | 0% |

| | Anti-ribosomal P-proteins ELISA (IgG)
Ratio | | |
|------------------|------------------------------------------------|-----------|-----------|
| | Sample 7 | Sample 8 | Sample 9 |
| n | 11** | 11** | 11** |
| Mean value (x): | 4.8 | 5.2 | 7.6 |
| Range of values: | 3.9 - 5.3 | 4.4 - 6.2 | 6.5 – 8.5 |
| Expected result: | positive | positive | positive |
| % positive: | 100% | 100% | 100% |
| % negative: | 0% | 0% | 0% |

*3 lots x 2 runs ** n lots x 1 run

• Linearity/assay reportable range: b. Not applicable.
• High dose Hook effect C.

Potential for a high dose Hook effect is a phenomenon that is inherent with one step "sandwich" assay designs: Very high concentrations of antigen in the patient sample bind to all available sites saturating them - on both the antibody-solid phase and the antibody-labeled conjugate and thereby prevent the "sandwich" formation. Under these conditions, the measured level of analyte may be significantly lower than the actual level present in the sample. The two-step immunoassay design of the Anti-ribosomal P-proteins ELISA (IgG) eliminates the adverse contribution of binding proteins, endogenous interfering substances and general matrix effects due to the extra wash step.

38

Image /page/38/Picture/1 description: The image shows a black and white pattern. The pattern consists of a series of black rectangles connected by black, rounded shapes. The rectangles are arranged horizontally, and the rounded shapes are positioned between them, creating a repeating sequence.

Traceability. Stability, Expected values (controls, calibrators or methods): · d.

A recognized standard or reference material for anti-nuclear antibodies is not available. Results of this assay are given in ratios. The reactivity of the Anti-ribosomal P-proteins ELISA was verified using the ANA reference panel of the CDC Centers for Disease Control and Prevention, Atlanta, USA.

• Limit of detection: ୧.
  • Not applicable.
• r. Analytical specificity:

Cross-reactivity: The quality of the antigen coated on the plates ensures a high specificity of the ELISA. Cross reactivity was investigated using a panel of 30 sera serologically positive for antibodies against nRNP/Sm, Sm, SS-A, Scl-70, Jo-1, centromeres, Rubella virus, Measles virus, Herpes simplex virus type 1 and Borrelia burgdorferi. All 30 sera were negative in the Anti-ribosomal Pproteins ELISA (IgG), so no cross reactivity is expected.

Interference: To investigate the influence from hemoglobin, triglycerides and bilirubin, 5 different sera at different anti-ribosomal P-proteins concentrations (13 - 104 RU/ml) were spiked with potential interfering substances and were incubated with the test system according to the package insert. Interferences from rheumatoid factor were investigated similarly by spiking with a RF positive serum. The recovery in relation to the unspiked sample without interferent was calculated. The individual recovery was within the range of 84 - 115 %. No significant interference was observed for concentrations of up to 1000 mg/dl for hemoglobin, 2000 mg/dl for triglyceride, 40 mg/dl for bilirubin and 500 IU/ml for rheumatoid factor.

• Assay cut-off: ರು
• Ratio 1.0
• 2. Comparison studies:
  • Method comparison with predicate device: a.

A comparison study was performed using 243 clinically characterized samples (90 SLE, 1 SLE/SS, 1 CLE, 51 Siögren's syndrome, 15 fibromyalgia, 35 RA, 30 borreliosis and 20 healthy), obtained from different sources from Europe and North America. The panel consisted of 49 men and 187 women. Age ranged from 14 to 83 years with an average age of 47 years. Anti-ribosomal P-proteins antibodies are expected in SLE. The other disease groups serve as control cohorts. The samples were tested with the EUROIMMUN Anti-ribosomal P-proteins ELISA (IgG) and with the Inova Quanta Lite Ribosomal P ELISA as the predicate device. All of the 10 discrepant samples were from patients with SLE, SLE/SS, and CLE.

| All samples

n = 243	Predicate ELISA
	positive	negative
EUROIMMUN
Anti-ribosomal P-proteins
ELISA (IgG)	28	10
negative	0	205
Negative agreement
Positive agreement
Overall agreement	$205 / 215 = 95.3%$
$28 / 28 = 100.0%$
$233 / 243 = 95.9%$	95% C.I.: 91.6% - 97.7%
95% C.I.: 87.7% - 100.0%
95% C.I.: 92.6% - 98.0%

• b. Matrix comparison:
  The usability of plasma was investigated using sample pairs each of serum and corresponding plasma (EDTA, Li-heparin, Citrate). Passing-Bablok regression was calculated for the comparison of serum to plasma. The regression equation is near the ideal correlation (intercept 0; slope 1.0) indicating equivalence of concentrations between serum and the corresponding plasma matrices. Coefficients of determination were found to be above 0.99 and %recovery compared toserum was in the range of 79 to 107 % (serum = 100 %).

	EDTA plasma	Li-heparin plasma	Citrate plasma
n	16	16	16
Regression equation
(y = plasma, x = serum)	$y = -0.98 + 1.02 x$	$y = 0.18 + 0.98 x$	$y = -1.79 + 0.99 x$
95% C.I. of intercept	-5.78 - 0.56	-2.42 - 3.26	-5.55 - 0.25
95% C.I. of slope	0.99 - 1.06	0.92 - 1.03	0.97 - 1.04
Coefficient of determination R2	0.9966	0.9939	0.9959
Mean %recovery	97 %	98 %	95 %
Range of %recovery	79 - 107 %	80 - 107 %	85 - 107 %

39

3. Clinical studies:

Clinical studies were performed in cooperation with different sites. In total 876 clinically characterized samples (376 from SLE patients and 500 from control groups) were investigated for anti-ribosomal Pproteins antibodies (IgG). With the EUROIMMUN Anti-ribosomal P-proteins ELISA (IgG) a prevalence of 5.3% (95% C.I.; 3.3 - 8.1%) was found in SLE with a specificity of 99.2% (95% C.I.: 98.0 - 99.8%). The results are shown in the table below. 95% C.I. are calculated by the exact method.

• Clinical sensitivity: a.

No.	Panel	n	Anti-ribosomal P-proteins ELISA (IgG)
			positive	%	95% C.I.
1	Systemic lupus erythematosus	376	20	5.3%	3.3 - 8.1%
No.	Panel	n	Anti-ribosomal P-proteins ELISA (IgG)
			negative	%	95% C.I.
2	Polymyositis/dermatomyositis	151	149	98.7%	95.3 - 99.8%
3	Rheumatoid arthritis	90	90	100.0%	96.0 - 100.0%
4	Systemic sclerosis	66	66	100.0%	94.6 - 100.0%
5	Sjögren's syndrome	55	54	98.2%	90.3 - 100.0%
6	Mixed connective tissue diseases	45	44	97.8%	88.2 - 99.9%
7	Other autoimmune diseases*	63	63	100.0%	94.3 - 100.0%
8	Borreliosis	30	30	100.0%	88.4 - 100.0%
	Total	500	496	99.2%	98.0 - 99.8%

b. Clinical specificity:

from the following groups: AIH (n = 8), Grave's disease (n = 12), Hashimoto (n = 11), celiac disease (n = 11), Diabetes Type I (n = 12)

• Other clinical supportive data (when a. and b. are not applicable): C. Not applicable.
• Clinical cut-off: See Assay cut-off.
• 5. Expected values/Reference range:

The levels of anti-ribosomal P-proteins antibodies (IgG) were analyzed in a panel of 150 samples from apparently healthy blood donors (79 men and 71 women with an average age of 38 y; age range: 18 – 67 y). The results are shown in the table below.

n	150
Positives	0
Negatives	150
Prevalence	0.0%
	Ratio
Lowest value	0.0
Highest value	0.7
Mean value	0.1
Std deviation	0.08

N. Proposed Labeling:

The labeling is sufficient and it satisfies the requirements of 21 CFR Part 809.10.

O. Conclusion:

The submitted information in this premarket notification is complete and supports a substantial equivalence decision.

Michael Locke
Signature

Dir. Regulatory Affairs Title

12 June 2013
Date

40

Image /page/40/Picture/0 description: The image shows the logo for the U.S. Department of Health & Human Services. The logo consists of a stylized eagle with three stripes forming its wing and body. The eagle is positioned to the right of the text, which is arranged in a circular pattern around the logo. The text reads "DEPARTMENT OF HEALTH & HUMAN SERVICES - USA".

DEPARTMENT OF HEALTH & HUMAN SERVICES

Public Health Service

Food and Drug Administration 10903 New Hampshire Avenue Document Control Center - WO66-G609 Silver Spring, MD 20993-0002

June 12, 2013

THE BINDING SITE GROUP LTD C/O MS. KATHRYN KOHL MANAGING DIRECTOR 1100 THE AMERICAN ROAD MORRIS PLAINS, NJ 07950

Re: K123261

Trade/Device Name: Euroimmun Anti-nRNP/Sm ELISA (IgG) Euroimmun Anti-Sm ELISA (IgG) Euroimmun Anti-SS-A ELISA (IgG) Euroimmun Anti-SS-B ELISA (IgG) Euroimmun Anti-Scl-70 ELISA (IgG) Euroimmun Anti-Centromeres ELISA (IgG) Euroimmun Anti-Jo-1 ELISA (IgG) Euroimmun Anti-Ribosomal P-Proteins ELISA (IgG) Regulation Number: 21 CFR 866.5110 Regulation Name: Antinuclear Antibody Immunological Test System Regulatory Class: II Product Code: LKO, LKP, LLL, LJM, MQA Dated: May 16, 2013 Received: May 17, 2013

Dear Ms. Kohl:

We have reviewed your Section 510(k) premarket notification of intent to market the device referenced above and have determined the device is substantially equivalent (for the indications for use stated in the enclosure) to legally marketed predicate devices marketed in interstate commerce prior to May 28, 1976, the enactment date of the Medical Device Amendments, or to devices that have been reclassified in accordance with the provisions of the Federal Food, Drug, and Cosmetic Act (Act) that do not require approval of a premarket approval application (PMA). You may, therefore, market the device, subject to the general controls provisions of the Act. The general controls provisions of the Act include requirements for annual registration, listing of devices, good manufacturing practice, labeling, and prohibitions against misbranding and adulteration. Please note: CDRH does not evaluate information related to contract liability warranties. We remind you, however, that device labeling must be truthful and not misleading.

If your device is classified (see above) into either class II (Special Controls) or class III (PMA), it may be subject to additional controls. Existing major regulations affecting your device can be found in the Code of Federal Regulations, Title 21, Parts 800 to 898. In addition, FDA may publish further announcements concerning your device in the Federal Register.

41

Please be advised that FDA's issuance of a substantial equivalence determination does not mean that FDA has made a determination that your device complies with other requirements of the Act or any Federal statutes and regulations administered by other Federal agencies. You must comply with all the Act's requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Parts 801 and 809); medical device reporting (reporting of medical device-related adverse events) (21 CFR 803); good manufacturing practice requirements as set forth in the quality systems (QS) regulation (21 CFR Part 820); and if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR 1000-1050.

If you desire specific advice for your device on our labeling regulations (21 CFR Parts 801 and 809), please contact the Division of Small Manufacturers, International and Consumer Assistance at its toll-free number [phone] or [phone] or at its Internet address http://www.fda.gov/MedicalDevices/ResourcesforYou/Industrv/default.htm. Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21CFR Part 807.97). For questions regarding the reporting of adverse events under the MDR regulation (21 CFR Part 803), please go to

http://www.fdai.gov/MedicalDevices/Safetv/ReportaProblem/default.htm for the CDRH's Office of Surveillance and Biometrics/Division of Postmarket Surveillance.

You may obtain other general information on your responsibilities under the Act from the Division of Small Manufacturers, International and Consumer Assistance at its toll-free number [phone] or [phone] or at its Internet address

http://www.fda.gov/MedicalDevices/ResourcesforYou/Industry/default.htm.

Sincerely yours.

Maria.M.Chan -S

Maria M. Chan, Ph.D. Director Division of Immunology and Hematology Devices Office of In Vitro Diagnostics and Radiological Health Center for Devices and Radiological Health

Enclosure

42

510(k) Number (if known): K123261

Device Name: Anti-ribosomal P-proteins ELISA (IgG)

Indications For Use:

The EUROIMMUN Anti-ribosomal P-proteins ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against ribosomal P-proteins in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of systemic lupus erythematosus, in conjunction with other laboratory and clinical findings.

Prescription Use × -(Part-21-CFR-801-Subpart-D)- AND/OR

Over-The-Counter Use -(21-CFR-807-Subpart-C)-

(PLEASE DO NOT WRITE BELOW THIS LINE-CONTINUE ON ANOTHER PAGE IF NEEDED)

Concurrence of CDRH; Office of In Vitro Diagnostics and Radiological Health (OIR)

Maria M. Chan -S

Division Sign-Off Office of In Vitro Diagnostics and Radiological Health

510(k): k 123261

43

510(k) Number (if known): K123261

Device Name: Anti-nRNP/Sm ELISA (IqG)

Indications For Use:

The EUROIMMUN Anti-nRNP/Sm ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against nRNP/Sm in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of mixed connective tissue diseases and systemic lupus erythematosus, in conjunction with other laboratory and clinical findings.

Prescription Use × (Part 21 CFR 801 Subpart D) AND/OR

Over-The-Counter Use (21 CFR 807 Subpart C)

(PLEASE DO NOT WRITE BELOW THIS LINE-CONTINUE ON ANOTHER PAGE IF NEEDED)

Concurrence of CDRH; Office of In Vitro Diagnostics and Radiological Health (OIR)

Maria M. Chan -S

Division Sign-Off Office of In Vitro Diagnostics and Radiological Health

510(k):

44

510(k) Number (if known): K123261

Device Name: Anti-Sm ELISA (IgG)

Indications For Use:

The EUROIMMUN Anti-Sm ELISA (IgG) test kit is intended for the gualitative determination of IgG class autoantibodies against Sm in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of systemic lupus erythematosus, in conjunction with other laboratory and clinical findings.

Prescription Use × (Part 21 CFR 801 Subpart D) AND/OR

Over-The-Counter Use (21 CFR 807 Subpart C)

(PLEASE DO NOT WRITE BELOW THIS LINE-CONTINUE ON ANOTHER PAGE IF NEEDED)

Concurrence of CDRH; Office of In Vitro Diagnostics and Radiological Health (OIR)

Maria M. Chan -S

Division Sign-Off Office of In Vitro Diagnostics and Radiological Health

45

510(k) Number (if known): K123261

Device Name: Anti-SS-A ELISA (IgG)

Indications For Use:

The EUROIMMUN Anti-SS-A ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against SS-A in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of Sjögren's syndrome and systemic lupus erythematosus, in conjunction with other laboratory and clinical findings.

Prescription Use × (Part 21 CFR 801 Subpart D) Over-The-Counter Use

(21 CFR 807 Subpart C)

(PLEASE DO NOT WRITE BELOW THIS LINE-CONTINUE ON ANOTHER PAGE IF NEEDED)

Concurrence of CDRH; Office of In Vitro Diagnostics and Radiological Health (OIR)

AND/OR

Maria M. Chan -S

Division Sign-Off Office of In Vitro Diagnostics and Radiological Health

510(k): K123261

46

510(k) Number (if known): K123261

Device Name: Anti-SS-B ELISA (IgG)

Indications For Use:

The EUROIMMUN Anti-SS-B ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against SS-B in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of Sjögren's syndrome and systemic lupus erythematosus, in conjunction with other laboratory and clinical findings.

Prescription Use × (Part 21 CFR 801 Subpart D) Over-The-Counter Use (21 CFR 807 Subpart C)

(PLEASE DO NOT WRITE BELOW THIS LINE-CONTINUE ON ANOTHER PAGE IF NEEDED)

Concurrence of CDRH; Office of In Vitro Diagnostics and Radiological Health (OIR)

AND/OR

Maria M. Chan -S

Division Sign-Off Office of In Vitro Diagnostics and Radiological Health

47

510(k) Number (if known): K123261

Device Name: Anti-Scl-70 ELISA (IgG)

Indications For Use:

The EUROIMMUN Anti-Scl-70 ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against Scl-70 in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of progressive systemic sclerosis, in conjunction with other laboratory and clinical findings.

Prescription Use × (Part 21 CFR 801 Subpart D) AND/OR

Over-The-Counter Use (21 CFR 807 Subpart C)

(PLEASE DO NOT WRITE BELOW THIS LINE-CONTINUE ON ANOTHER PAGE IF NEEDED)

Concurrence of CDRH; Office of In Vitro Diagnostics and Radiological Health (OIR)

Maria M. Chan -S

Division Sign-Off Office of In Vitro Diagnostics and Radiological Health

48

510(k) Number (if known): K123261

Device Name: Anti-Centromeres ELISA (IgG)

Indications For Use:

The EUROIMMUN Anti-Centromeres ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against centromeres in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of limited form of progressive systemic sclerosis (CREST syndrome), in conjunction with other laboratory and clinical findings.

Prescription Use X (Part 21 CFR 801 Subpart D) Over-The-Counter Use (21 CFR 807 Subpart C)

(PLEASE DO NOT WRITE BELOW THIS LINE-CONTINUE ON ANOTHER PAGE IF NEEDED)

Concurrence of CDRH; Office of In Vitro Diagnostics and Radiological Health (OIR)

AND/OR

Maria M. Chan -S

Division Sign-Off Office of In Vitro Diagnostics and Radiological Health

49

510(k) Number (if known): K123261

Device Name: Anti-Jo-1 ELISA (IgG)

Indications For Use:

The EUROIMMUN Anti-Jo-1 ELISA (IgG) test kit is intended for the qualitative determination of IgG class autoantibodies against Jo-1 in human serum and plasma (EDTA, Li-heparin, Citrate). It is used as an aid in the diagnosis of polymyositis and dermatomyositis, in conjunction with other laboratory and clinical findings.

Prescription Use × (Part 21 CFR 801 Subpart D) AND/OR

Over-The-Counter Use (21 CFR 807 Subpart C)

(PLEASE DO NOT WRITE BELOW THIS LINE-CONTINUE ON ANOTHER PAGE IF NEEDED)

Concurrence of CDRH; Office of In Vitro Diagnostics and Radiological Health (OIR)

Maria M. Chan -S

Division Sign-Off Office of In Vitro Diagnostics and Radiological Health

510(k): K123261