← Product Code [MID](/submissions/IM/subpart-f%E2%80%94immunological-test-systems/MID) · K091845

# CARDIOLIPIN AUTOANTIBODY IMMUNOLOGICAL TEST SYSTEM (K091845)

_Phadia US, Inc. · MID · Jun 7, 2010 · Immunology · SESE_

**Canonical URL:** https://fda.innolitics.com/submissions/IM/subpart-f%E2%80%94immunological-test-systems/MID/K091845

## Device Facts

- **Applicant:** Phadia US, Inc.
- **Product Code:** [MID](/submissions/IM/subpart-f%E2%80%94immunological-test-systems/MID.md)
- **Decision Date:** Jun 7, 2010
- **Decision:** SESE
- **Submission Type:** Traditional
- **Regulation:** 21 CFR 866.5660
- **Device Class:** Class 2
- **Review Panel:** Immunology

## Indications for Use

EliA Cardiolipin IgG is intended for the in vitro semi-quantitative measurement of IgG antibodies directed to cardiolipin in human serum and plasma (Li-heparin, EDTA, citrate) to aid in the diagnosis of antiphospholipid syndrome (APS) as well as thrombotic disorders related to secondary antiphospholipid syndrome in conjunction with other laboratory and clinical findings. EliA Cardiolipin IgG uses the the EliA IgG method on the instruments Phadia 100 and Phadia 250. EliA Cardiolipin IgM is intended for the in vitro semi-quantitative measurement of IgM antibodies directed to cardiolipin in human serum and plasma (Li-heparin, EDTA, citrate) to aid in the diagnosis of antiphospholipid syndrome (APS) as well as thrombotic disorders related to secondary antiphospholipid syndrome in conjunction with other laboratory and clinical findings. EliA Cardiolipin IgM uses the the EliA IgM method on the instruments Phadia 100 and Phadia 250. EliA B2-Glycoprotein I IgG is intended for the in vitro semi-quantitative measurement of IgG antibodies directed to B2-Glycoprotein I in human serum and plasma (Li-heparin, EDTA, citrate) to aid in the diagnosis of antiphospholipid syndrome (APS) a as well as thrombotic disorders related to secondary antiphospholipid syndrome in conjunction with other laboratory and clinical findings. EliA B2-Glycoprotein I IgG uses the the EliA IgG method on the instruments Phadia 100 and Phadia 250. EliA B2-Glycoprotein I IgM is intended for the in vitto semi-quantitative measurement of IgM antibodies directed to ß2-Glycoprotein I in human serum and plasma (Li-heparin, EDTA, citrate) to aid in the diagnosis of antiphospholipid syndrome (APS) as well as thrombotic disorders related to secondary antiphospholipid syndrome in conjunction with other laboratory and clinical findings. EliA B2-Glycoprotein I IgM uses the the EliA IgM method on the instruments Phadia 100 and Phadia 250. EliA APS Positive Control 100 is intended for laboratory use in monitoring the performance of in vitro measurement of antibodies to cardiolipin and B2-Glycoprotein I with Phadia 100 using the EliA IgG or IgM method. EliA APS Positive Control 250 is intended for laboratory use in monitoring the performance of in vitro measurement of antibodies to cardiolipin and ß2-Glycoprotein I with Phadia 250 using the EliA IgG or IgM method.

## Device Story

Fully automated fluorescence-immunoassay system; utilizes EliA single wells as solid phase; performed on Phadia 100/250 instruments. Input: human serum/plasma samples. Principle: non-competitive solid phase ELISA; patient antibodies bind to antigen-coated wells; enzyme-labeled conjugate (mouse anti-human IgG/IgM beta-galactosidase) added; fluorescence measured after development solution reaction. Output: semi-quantitative antibody levels. Used in clinical laboratories; operated by trained personnel. Results aid physicians in diagnosing APS and related thrombotic disorders by comparing patient response to standardized calibrators. Benefits: standardized, automated assessment of autoimmune markers.

## Clinical Evidence

Comparison study between new and predicate devices, testing of clinically defined sera, and testing of samples from apparently healthy subjects.

## Technological Characteristics

Semi-quantitative immunofluorescence assay; bovine cardiolipin/β2-GPI or human β2-GPI antigen-coated microwells; β-galactosidase labeled mouse monoclonal anti-human IgG/IgM conjugate; automated Phadia 100/250 instrumentation; 37°C controlled reaction; fluorescence signal detection; PBS-based diluents with 0.095% sodium azide.

## Regulatory Identification

A multiple autoantibodies immunological test system is a device that consists of the reagents used to measure by immunochemical techniques the autoantibodies (antibodies produced against the body's own tissues) in serum and other body fluids. Measurement of multiple autoantibodies aids in the diagnosis of autoimmune disorders (disease produced when the body's own tissues are injured by autoantibodies).

## Predicate Devices

- Varelisa Cardiolipin IgG Antibodies (k020752)
- Varelisa Cardiolipin IgM Antibodies (k020758)
- Varelisa β2-Glycoprotein I IgG Antibodies (k040449)
- Varelisa β2-Glycoprotein I IgM Antibodies (k040451)

## Submission Summary (Full Text)

> This content was OCRed from public FDA records by [Innolitics](https://innolitics.com). If you use, quote, summarize, crawl, or train on this content, cite Innolitics at https://innolitics.com.
>
> Innolitics is a medical-device software consultancy. We help companies design, build, and clear FDA-regulated software and AI/ML devices, including [a 510(k)](https://innolitics.com/services/510ks/), [a De Novo](https://innolitics.com/services/regulatory/), [a SaMD](https://innolitics.com/services/end-to-end-samd/), [an AI/ML medical device](https://innolitics.com/services/medical-imaging-ai-development/), or [an FDA regulatory strategy](https://innolitics.com/services/regulatory/).

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# 510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION DECISION SUMMARY

A. 510(k) Number:
k091845

B. Purpose for Submission:
New device

C. Measurand:
IgG and IgM Antibodies to cardiolipin
IgG and IgM Antibodies to β2-glycoprotein I

D. Type of Test:
Semi-quantitative immunofluorescence assay

E. Applicant:
Phadia US Inc.

F. Proprietary and Established Names:
EliA™ Cardiolipin IgG Immunoassay
EliA™ Cardiolipin IgM Immunoassay
EliA™ β2-Glycoprotein I IgG Immunoassay
EliA™ β2-Glycoprotein I IgM Immunoassay
EliA™ APS Positive Control 100
EliA™ APS Positive Control 250

G. Regulatory Information:

1. Regulation section:
21 CFR § 866.5660, Multiple Autoantibodies Immunological Test System
21 CFR § 866.5660, Multiple Autoantibodies Immunological Test System
21 CFR § 862.1660- Quality Control Material (assayed and unassayed)

2. Classification:
Class II (Assays)
Class I (Controls)

3. Product code:
MID, System Test, Anti-cardiolipin Immunological
MSV, Antibodies, β2- Glycoprotein I (β2-GPI)
JJY, A quality control material (assayed and unassayed)

4. Panel:
Immunology (82) (Assays)
Chemistry (75) (Controls)

H. Intended Use:

1. Intended use(s):
EliA Cardiolipin IgG is intended for the in vitro semi-quantitative measurement of IgG antibodies directed to cardiolipin in human serum and plasma (Li-heparin, EDTA, citrate) to aid in the diagnosis of antiphospholipid syndrome (APS) as well as thrombotic disorders related to secondary antiphospholipid syndrome in conjunction with other laboratory and clinical findings. EliA Cardiolipin IgG uses the EliA IgG method on the instrument Phadia100 and Phadia 250.

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EliA Cardiolipin IgM is intended for the in vitro semi-quantitative measurement of IgM antibodies directed to cardiolipin in human serum and plasma (Li-heparin, EDTA, citrate) to aid in the diagnosis of antiphospholipid syndrome (APS) as well as thrombotic disorders related to secondary antiphospholipid syndrome in conjunction with other laboratory and clinical findings. EliA Cardiolipin IgM uses the EliA IgM method on the instruments Phadia 100 and Phadia 250.

EliA β2-Glycoprotein I IgG is intended for the in vitro semi-quantitative measurement of IgG antibodies directed to β2-Glycoprotein I in human serum and plasma (Li-heparin, EDTA, citrate) to aid in the diagnosis of antiphospholipid syndrome (APS) as well as thrombotic disorders related to secondary antiphospholipid syndrome in conjunction with other laboratory and clinical findings. EliA β2-Glycoprotein I IgG uses the EliA IgG method on the instruments Phadia® 100 and Phadia® 250.

EliA β2-Glycoprotein I IgM is intended for the in vitro semi-quantitative measurement of IgM antibodies directed to β2-Glycoprotein I in human serum and plasma (Li-heparin, EDTA, citrate) to aid in the diagnosis of antiphospholipid syndrome (APS) as well as thrombotic disorders related to secondary antiphospholipid syndrome in conjunction with other laboratory and clinical findings. EliA β2-Glycoprotein I IgM uses the EliA IgM method on the instruments Phadia 100 and Phadia 250.

EliA™ APS Positive Control 100 is intended for laboratory use in monitoring the performance of in vitro measurement of antibodies to cardiolipin and β2-Glycoprotein I with Phadia 100 using the EliA IgG or IgM method.

EliA™ APS Positive Control 250 is intended for laboratory use in monitoring the performance of in vitro measurement of antibodies to cardiolipin and β2-Glycoprotein I with Phadia 250 using the EliA IgG or IgM method.

2. Indication(s) for use:
Same as intended use

3. Special conditions for use statement(s):
For prescription use only

4. Special instrument requirements:
Phadia® 100 and Phadia® 250 (k061165)

I. Device Description:
The method specific reagents on ImmunoCAP 100 and ImmunoCAP 250 are identical; they are only filled in different containers.

Each device consists of: 1) antigen coated wells for 48 determinations (bovine cardiolipin antigen and bovine β2-glycoprotein I as co-factor or β2-glycoprotein I antigen) - 4 carriers (12 wells each), ready to use; 2) EliA method-specific sample diluent, PBS with 0.095% sodium azide - 6 vials, 9 mL each, ready to use; 3) EliA IgG/IgM conjugate, β-galactosidase labeled anti IgG/IgM (mouse monoclonal antibodies) - 2 or 6 vials, 4.8 mL

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each, ready to use; 4) positive control: human monoclonal IgG and IgM antibodies specific to cardiolipin and monoclonal IgG and IgM antibodies specific to  $\beta 2$ -Glycoprotein I - 6 single use vials(0.3 mL each), ready to use; 5) negative control: containing normal human serum from healthy donors - 6 single-use vials, 0.3 mL each, ready to use; 6) EliA calibrators, human IgG/IgM in PBS - 6 single-use vials, 0.3 mL each, ready to use. The "Phadia EliA Immunodiagnostic System" is automated system for immunodiagnostic testing. The test specific, method specific and general reagents are packaged and purchased as separate units.

# J. Substantial Equivalence Information:

1. Predicate device name(s) and 510(k) number(s):

Varelisa Cardiolipin IgG Antibodies (k020752)

Varelisa Cardiolipin IgM Antibodies (k020758)

Varelisa  $\beta 2$ -Glycoprotein I IgG Antibodies (k040449)

Varelisa  $\beta 2$ -Glycoprotein I IgM Antibodies (k040451)

2. Comparison with predicate:

EliATM Cardiolipin IgG vs.Varelisa Cardiolipin IgG Antibodies

EliATM Cardiolipin IgM vs. Varelisa Cardiolipin IgM Antibodies

EliA  $\beta 2$ -GP I IgG Well vs. Varelisa  $\beta 2$ -Glycoprotein I IgG Antibodies

EliATM  $\beta 2$ -Glycoprotein I IgM vs. Varelisa  $\beta 2$ -Glycoprotein I IgM Antibodies

|  Similarities  |   |   |
| --- | --- | --- |
|  Item | New EliA™ Device | Predicate Varelisa Device  |
|  Intended Use | EliA™ Cardiolipin IgG/IgM and β2-Glycoprotein I IgG/IgM are intended for the in vitro semi-quantitative measurement of IgG/IgM antibodies directed to cardiolipin or β2-Glycoprotein I in serum and plasma (heparin, EDTA, citrate) to aid in the diagnosis of antiphospholipid syndrome (APS) as well as thrombotic disorders related to secondary antiphospholipid syndrome in conjunction with other laboratory and clinical findings. EliA β2-Glycoprotein I IgG uses the EliA IgG method on the instruments Phadia 100 and Phadia 250. | The Varelisa Cardiolipin IgG/IgM and β2-Glycoprotein I IgG/IgM Antibodies EIA kits are designed for the semi-quantitative and qualitative determination of IgG/IgM antibodies against cardiolipin or β2-Glycoprotein I in serum or plasma to aid in the diagnosis of antiphospholipid syndrome (APS) and to evaluate the thrombotic risk in patients with systemic lupus erythematosus (SLE).  |
|  Controls | Positive and Negative Control Sera provided with the EliA APS | Same  |
|  Assay Type | ELISA | Same  |
|  Type of test | Semi-quantitative | Qualitative and semi-quantitative  |
|  Antigen used | Bovine cardiolipin antigen and bovine β2-glycoprotein as co-factor or Human purified β2-Glycoprotein I | Same  |

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|  Similarities  |   |   |
| --- | --- | --- |
|  Item | New EliATM Device | Predicate Varelisa Device  |
|  Solid Phase | Microwells | Same  |
|  Differences  |   |   |
| --- | --- | --- |
|  Item | Device | Predicate  |
|  Concept | Modular reagents concept (test-method specific and general reagents) | All reagents in a single kit  |
|  Instrumentation | Phadia 100 and Phadia 250 are fully automated immunoassay analyzers | ELISA-Reader  |
|  Reaction temperature | 37°C controlled | Room temperature, 20-26°C  |
|  Detection antibody (conjugate) | mouse anti-human IgG/IgM β-Galactosidase labeled monoclonal antibodies | goat anti-human IgG/IgM horse-radish peroxidase labeled  |
|  Signal | Fluorescence | Optical density  |
|  Calibration | Total IgG and IgM Calibration | Analyte specific IgG Calibration  |
|  Calibration curve | Option to store curve for up to 28 days and run curve controls in each assay for calibration | Calibration Curve in each assay  |
|  Calibrators: IgG Anti-Cardiolipin and anti-β2-glycoprotein I | 0, 4, 10, 20, 100, 600 μg/L | 0, 4, 8, 20, 50, 100 U/mL  |
|  IgM Anti-Cardiolipin and anti-β2-glycoprotein I | 0,10,35,80,500,1000 μg/L | 0, 4, 8, 20, 50, 100 U/mL  |
|  Cardiolipin Measuring range | IgG 4.4-418 GPL-U/mL IgM 2.5 - 466 MPL-U/mL | 1-100 GPL-U/mL 1-100 MPL-U/mL  |
|  Cardiolipin IgG/IgM Cut-off | Negative - < 10 GPL-U/mL Weak positive - 10-40 GPL-U/mL Positive - > 40 GPL-U/mL | Negative - < 10 GPL-U/mL Equivocal - 10-15 GPL-U/mL Positive - > 15 GPL-U/mL  |
|  β2-Glycoprotein I measuring range | IgG 3.8 - 532 U/mL IgM 3.9 - 500 U/mL | 1.0 - 100 U/mL 1.0 - 100 U/mL  |
|  β2-Glycoprotein I Cut-off | Negative - < 7 U/mL Equivocal - 7-10 U/mL Positive - > 10 U/mL | Negative - < 10 U/mL Equivocal - 10-15 U/mL Positive - > 15 U/mL  |

K. Standard/Guidance Document Referenced (if applicable):
Not applicable

L. Test Principle:

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The EliA test wells are coated with bovine cardiolipin antigen and bovine  $\beta 2$ -glycoprotein I as co-factor or with human  $\beta 2$ -Glycoprotein I. If present in the patient's specimen, antibodies to cardiolipin or  $\beta 2$ -glycoprotein I bind to their specific antigen. After washing away non-bound antibodies, enzyme-labeled antibodies against human IgG or IgM antibodies, are added to form an antibody-conjugate complex (EliA  $\beta$ -Galactosidase anti - IgG/IgM mouse monoclonal antibodies). After incubation, non-bound conjugate is washed away and the bound complex is incubated with a Development Solution. After stopping the reaction, the fluorescence in the reaction mixture is measured. The higher the value, the more IgG or IgM is present in the specimen. To evaluate test results, the signal response for patient samples is compared directly to the signal response for calibrators of defined value.

# M. Performance Characteristics (if/when applicable):

# 1. Analytical performance:

# a. Precision/Reproducibility:

To determine the precision of the assays on the Phadia 100 and Phadia 250 instrument, the variability was assessed on 6 samples. Each sample was run on 3 instruments x 6 runs each over 6 days (equal to 108 replicate determinations per sample) with a calibration curve in each run. Additional 2-4 samples were run on the analytes that showed  $&gt;10\%$  CV. The statistical evaluation was performed by Analysis of Variance. The results are summarized in the tables below:

|  EliA Cardiolipin IgG IC100  |   |   |   |
| --- | --- | --- | --- |
|  Mean value GPL - U/mL | Coefficients of variation (%)  |   |   |
|   |  Intra Run CV% | Inter Run (between days) CV% | Total Imprecision CV%  |
|  7.8 | 3.7 | 2.4 | 4.4  |
|  14.0* | 5.5* | 3.1* | 6.4*  |
|  16.6 | 10.2 | 2.1 | 10.4  |
|  47.3 | 4.1 | 2.2 | 4.6  |
|  82.9 | 3.9 | 2.2 | 4.5  |
|  164.5 | 8.7 | 4.2 | 9.7  |
|  166.7* | 5.1* | 4.6* | 6.8*  |
|  188.2* | 4.3* | 3.4* | 5.5*  |
|  443.8 | 8.5 | 5.1 | 9.9  |
|  EliA Cardiolipin IgG IC250  |   |   |   |
| --- | --- | --- | --- |
|  Mean value GPL - U/mL | Coefficients of variation (%)  |   |   |
|   |  Intra Run CV% | Inter Run (between days) CV% | Total Imprecision CV%  |
|  8.0 | 3.9 | 2.8 | 4.8  |
|  16.8 | 8.9 | 0.3 | 8.9  |
|  18.2* | 3.4* | 3.4* | 4.8*  |
|  30.4* | 6.5* | 3.4* | 7.3*  |
|  47.2 | 3.1 | 2.4 | 3.9  |
|  80.3 | 3.8 | 2.8 | 4.7  |
|  178.9 | 17.3 | 8.7 | 19.4  |
|  239.8* | 3.3* | 2.9* | 4.4*  |
|  377.1* | 5.6* | 4.3* | 7.0*  |
|  418.6 | 6.2 | 5.2 | 8.1  |

* Additional samples tested for analytes with &gt;10% CV

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EliA Cardiolipin IgM IC100

|  Mean value MPL-U/mL | Coefficients of variation (%)  |   |   |
| --- | --- | --- | --- |
|   |  Intra Run CV% | Inter Run (between days) CV% | Total Imprecision CV%  |
|  3.3 | 9.3 | 9.6 | 13.4  |
|  21.9 | 4.9 | 5.6 | 7.5  |
|  52.5 | 4.1 | 4.7 | 6.2  |
|  105.3 | 3.7 | 5.2 | 6.4  |
|  184.7 | 6.8 | 5.7 | 8.9  |
|  420.2 | 4.5 | 2.1 | 5.0  |

EliA Cardiolipin IgM IC250

|  Mean value MPL - U/mL | Coefficients of variation (%)  |   |   |
| --- | --- | --- | --- |
|   |  Intra Run CV% | Inter Run (between days) CV% | Total Imprecision CV%Cv  |
|  2.0 | 12.0 | 8.9 | 15.0  |
|  24.0 | 3.3 | 5.4 | 6.3  |
|  53.0 | 3.7 | 3.2 | 4.9  |
|  105.1 | 3.5 | 3.2 | 4.7  |
|  210.1 | 5.0 | 3.7 | 6.2  |
|  490.8 | 3.1 | 1.6 | 3.5  |

EliA b2-Glycoprotein I IgG IC100

|  Mean value U/mL | Coefficients of variation (%)  |   |   |
| --- | --- | --- | --- |
|   |  Intra Run CV% | Inter Run (between days) CV% | Total Imprecision CV%  |
|  4.6 | 6.1 | 6.0 | 8.6  |
|  8.7 | 7.2 | 15.5 | 17.1  |
|  9.0* | 5* | 4.4* | 6.7*  |
|  10.9* | 3.7* | 3.9* | 5.3*  |
|  13.9 | 5.1 | 3.7 | 6.3  |
|  39.8 | 3.8 | 2.3 | 4.5  |
|  113.7 | 4.4 | 4.8 | 6.5  |
|  271.1 | 4.7 | 3.5 | 5.9  |
|  501.0 | 7.2 | 4.9 | 8.7  |

EliA b2-Glycoprotein I IgG IC250

|  Mean value U/mL | Coefficients of variation (%)  |   |   |
| --- | --- | --- | --- |
|   |  Intra Run CV% | Inter Run (between days) CV% | Total Imprecision CV%  |
|  4.4 | 3.4 | 4.3 | 5.5  |
|  9.1* | 5.3* | 3.8* | 6.5*  |
|  10.7 | 7.8 | 12.9 | 15.1  |
|  14.1* | 4.6* | 3.9* | 6.0*  |
|  15.5 | 4.7 | 3.8 | 6.0  |
|  43.0 | 3.7 | 3.0 | 4.8  |
|  112.1 | 4.5 | 3.1 | 5.5  |
|  231.0 | 3.3 | 2.8 | 4.3  |
|  449.3 | 4.4 | 3.8 | 5.8  |

* Additional samples tested for analytes with &gt;10% CV

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EliA b2-Glycoprotein I IgM IC100

|  Mean value U/mL | Coefficients of variation (%)  |   |   |
| --- | --- | --- | --- |
|   |  Intra Run CV% | Inter Run (between days) CV% | Total Imprecision CV%  |
|  4.5 | 12.2 | 9.5 | 15.4  |
|  9.4 | 6.6 | 6.4 | 9.2  |
|  17.7 | 4.4 | 5.7 | 7.2  |
|  52.0 | 4.4 | 5.2 | 6.8  |
|  186.5 | 4.3 | 5.4 | 6.9  |
|  498.1 | 3.7 | 5.2 | 6.4  |

EliA b2-Glycoprotein I IgM IC250

|  Mean value U/mL | Coefficients of variation (%)  |   |   |
| --- | --- | --- | --- |
|   |  Intra Run CV% | Inter Run (between days) CV% | Total Imprecision CV%  |
|  3.3 | 11.5 | 14.7 | 18.7  |
|  8.9 | 4.2 | 8.2 | 9.2  |
|  18.2 | 2.7 | 1.9 | 3.3  |
|  58.6 | 4.3 | 3.0 | 5.2  |
|  196.5 | 4.0 | 3.9 | 5.5  |
|  433.8 | 3.0 | 4.3 | 5.2  |

b. Linearity/assay reportable range:

Nine samples per analyte were diluted according to the following scheme after the method specific dilution (1:10 for EliA Cardiolipin IgG and IgM assays and the EliA β2-Glycoprotein I IgG assay or 1:50 for the EliA β2-Glycoprotein I IgM assay): 1/2, 1/4, 1/8, 1/16 and 1/32 using EliA™ Sample Diluent. Each sample was run in triplicates in two runs. For each analyte, 8 of the 9 sample sets were assayed on the IC100 and one sample set on the IC250. The sample concentrations tested for linearity were as follows: Cardiolipin IgG - 141.2 to 568.9 GPL-U/mL; Cardiolipin - IgM 87.5 to 466.4; β2-Glycoprotein I IgG 128.9 to 806.4 U/mL; β2-Glycoprotein I IgM - 172.7 to 500.5 U/mL. Representative results are summarized in the tables below.

EliA™ Cardiolipin IgG

|  Instrument | EliA Cardiolipin IgG Dilution Range (GPL-U/mL) | Slope (95% CI) | Y-intercept (95% CI) | R² | %CV Range  |
| --- | --- | --- | --- | --- | --- |
|  IC100 | 4.4 - 141.2 | 1.00 (0.93 to 1.07) | -2.92 (-7.76 to 1.93) | 0.997 | 1.3 – 5.2  |
|   |  6.5 – 517.4 | 1.03 (1.00 to 1.06) | -12.34 (-17.90 to -6.77) | 0.999 | 0.1 – 2.2  |
|  IC250 | 6.0 – 568.9 | 1.00 (1.00 to 1.07) | -2.87 (-8.19 to 2.45) | 0.998 | 0.3 – 2.7  |

Claimed linearity range is 4.4-418 GPL U/mL.

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EliA™ Cardiolipin IgM

|  Instrument | EliA Cardiolipin IgM Dilution Range (MPL-U/mL) | Slope (95% CI) | Y-intercept (95% CI) | R² | %CV range  |
| --- | --- | --- | --- | --- | --- |
|  IC100 | 2.5 - 88.9 | 1.00 (0.99 to 1.01) | 0.32 (-0.10 to 0.73) | 1.000 | 1.3 – 6.0  |
|   |  7.2 – 399.0 | 1.01 (0.97 to 1.05) | -13.93 (-20.11 to -7.75) | 0.998 | 1.2 – 7.5  |
|  IC250 | 9.2 – 466.4 | 1.01 (0.98 to 1.05) | -17.90 (-24.86 to 10.94) | 0.998 | 0.4 – 3.1  |

Claimed linearity range is 2.5-466 MPL-U/mL.

EliA™ β2-Glycoprotein I IgG

|  Instrument | EliA β2-GPI IgG Dilution Range (U/mL) | Slope (95% CI) | Y-intercept (95% CI) | R² | %CV range  |
| --- | --- | --- | --- | --- | --- |
|  IC100 | 3.8 - 128.9 | 0.98 (0.80 to 1.15) | -4.10 (-15.79 to 7.58) | 0.983 | 0.5 – 5.7  |
|   |  9.4 – 742.3 | 1.04 (1.01 to 1.07) | -14.85 (-23.62 to -6.08) | 0.998 | 0.3 – 4.4  |
|  IC250 | 8.1 – 806.4 | 1.04 (0.99 to 1.09) | -4.50 (-20.34 to 11.35) | 0.995 | 0.3 – 4.4  |

Claimed linearity range is 3.8-532 U/mL.

EliA™ β2-Glycoprotein I IgM

|  Instrument | EliA β2-GPI IgM Dilution Range (U/mL) | Slope (95% CI) | Y-intercept (95% CI) | R² | %CV Range  |
| --- | --- | --- | --- | --- | --- |
|  IC100 | 3.9 to 176.8 | 0.99 (0.94 to 1.04) | 3.34 (-0.82 to 7.51) | 0.999 | 0.7 – 2.9  |
|   |  5.5 – 500.5 | 1.00 (0.97 to 1.04) | -14.04 (-21.14 to -6.93) | 0.998 | 1.2 – 14.1  |
|   |  5.5 – 172.7 | 0.98 (0.96 to 1.00) | 2.16 (0.37 to 3.95) | 0.999 | 1.2 – 14.1*  |
|  IC250 | 6.3 – 466.3 | 1.01 (0.97 to 1.04) | -14.25 (-20.52 to -7.97) | 0.998 | 0.8 – 5.6  |

Claimed linearity range is 3.9 - 500 U/mL.

Results above the upper limit are reported as "above". No recommendations are made for dilution of samples outside measuring range in the package insert.

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Values below detection limit are reported as &lt; x.x GPL-, MPL or EliA U/mL depending on the analyte.

High Dose Hook Effect:

IgG method: High dose hook effect was analyzed by using dilutions from high positive serum samples with an estimated concentration higher than the calibrator 600 (containing 600 µg/L of IgG). The sample dilutions were measured in 4 replicates.

EliA Cardiolipin IgG: A hook effect was not observed when analyzing a high positive sample that had a concentration up to 6.5 times above the upper limit of the measuring range. The upper limit of the measuring range was set to 418 GPL-U/mL.

EliA β2-Glycoprotein I Ig: A hook effect was not observed when analyzing a high positive sample that had a concentration up to 6.4 times above the upper limit of the measuring range. The upper limit of the measuring range was set to 532 U/mL.

IgM method: High dose hook effect was analyzed by using dilutions from high positive serum samples with an estimated concentration higher than the calibrator 1000 (containing 1000 µg/L of IgM). The sample dilutions were measured in 4 replicates.

EliA Cardiolipin IgM: A hook effect was not observed when analyzing a high positive sample that had a concentration up to 5.85 times above the upper limit of the measuring range. The upper limit of the measuring range was set to 466 MPL-U/mL.

EliA β2-Glycoprotein I IgM: A hook effect was not observed when analyzing a high positive sample that had a concentration up to 8.1 times above the upper limit of the measuring range. The upper limit of the measuring range was set to 500 U/mL.

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

The EliA IgG system reagents and EliA Negative Control are cleared under the k061165 for the EliA IgG system; k072149/k072393 for EliA Negative Control.

The IgG and IgM calibrators are traceable (via an unbroken chain of calibrations) to the International Reference Preparation (IRP) 67/86 of Human Serum Immunoglobulins A, G and M from WHO. New batches of IgG and IgM Calibrators are compared to a secondary standard (standardized with the IRP) or the IRP directly and adjusted accordingly to meet the correct concentration.

The instrument measures specific IgG/IgM concentrations in µg/L. By using a conversion factor given by the lot-specific code of the EliA Cardiolipin IgG and IgM Well, the results are automatically converted to GPL-U/mL or MPL-U/mL, respectively.

9

{9}

Shelf Life of test components:

Three batches of EliA Cardiolipin IgG/IgM or β2-Glycoprotein I IgG/IgM wells were tested in real time stability studies to support the claimed shelf life at 2-8°C up to 18 months.

EliA APS Positive Control stability:

Shelf of the EliA APS positive control is summarized below:

Open stability:

Open stability is not tested as the EliA APS Positive Control and Negative Control are packaged in single use vials.

Closed stability:

A study was done to investigate the accelerated stability of the EliA APS Positive Control. A preliminary shelf life of 18 months was given for the EliA APS Positive Control. Real time studies are being done alongside the accelerated studies.

d. Detection limit:

The purpose of this study was to verify the lower limit of the measuring ranges for EliA Cardiolipin IgG and EliA β2-Glycoprotein I IgG (detection limit) by providing the ability of the new device to differentiate between the background sample diluent) and a 1/8 dilution of the lowest calibrator point.

Detection limit (LoD) IgG method:

Calibrator 4.0 (containing 4 μg/L of IgG) was diluted 1:2 (i.e. 2 μg/L), 1:4 (i.e. 1 μg/L) and 1:8 (i.e. 0.5 μg/L) in EliA sample diluent and run on EliA IgG Calibrator wells. EliA Sample diluent was run on the different EliA Cardiolipin IgG and EliA β2-Glycoprotein I IgG wells. EliA sample diluent results were compared with the Calibrator results.

Detection limit (LoD) IgM method:

Calibrator 10.0 (containing 10 μg/L of IgM) was diluted 1:2 (i.e. 5 μg/L), 1:4 (i.e. 2.5 μg/L) and 1:8 (i.e. 1.25 μg/L) in EliA sample diluent and run on EliA IgM Calibrator wells. EliA Sample diluent was run on the different EliA Cardiolipin IgM and EliA β2-Glycoprotein I IgM wells. EliA sample diluent results were compared with the Calibrator results.

Results are summarized below:

EliA Cardiolipin IgG: LoD was set to 0.5 GPL-U/mL.

EliA β2-Glycoprotein I IgG: LoD was set to 0.6 U/mL.

EliA Cardiolipin IgM: LoD was set to 0.8 MPL-U/mL.

EliA β2-Glycoprotein I IgM: LoD was set to 0.9 U/mL.

f. Analytical specificity/Interference:

The purpose of this study was to investigate whether high concentrations of potentially interfering substances in serum, like bilirubin, hemoglobin, chyle and rheumatoid factor adversely affect the results of EliA assays. Two positive serum samples with concentration levels around the cut-off and one high positive serum sample with concentration level around calibrator 5 were spiked with the interfering substances bilirubin C, bilirubin F, hemoglobin, chyle or rheumatoid

10

{10}

factor. The same samples were also spiked with substance specific blanks. The samples were tested in 3 replicates. A calibration curve in two replicates was run in each assay. The runs were repeated twice. The table below shows the end concentration of the interference substances spiked into the 1:10 diluted serum samples:

|  Substance | Concentration in the 1:10 Diluted Serum sample  |
| --- | --- |
|  Bilirubin C | 20.6 μg/mL  |
|  Bilirubin F | 21.1 μg/mL  |
|  Hemoglobin | 519 μg/mL  |
|  Chyle | 1570 Units/dl  |
|  Rheumatoid factor | 55 IU/mL*  |

Bilirubin C, bilirubin F, chyle and hemoglobin did not adversely affect the results.

*Increased levels of rheumatoid factor may have an effect on the response on EliA Cardiolipin IgM Well and can influence the β2-Glycoprotein I IgM test result when the patient sample contains β2-Glycoprotein I IgG antibodies.

Generally, the use of sera containing lipemic, hemolized or microbial contaminations is not recommended as stated in the package inserts.

## Carry over study:

A sample with a specific anti-Cardiolipin IgM titer above the measuring range was first pipetted on EliA Dummy wells (wells which are not coated with Cardiolipin antigen). The pipette then was washed by the ordinary washing routine of the instrument. Thereafter pure diluent was pipetted on EliA Cardiolipin IgM wells. If there remained any of the sample with high specific anti-Cardiolipin IgM titer on the pipette surface it would increase the signal compared to the basic signal normally obtained when measuring pure diluent on EliA Cardiolipin IgM wells. This was repeated five times. The study has been performed on three instruments. Signal level for pure diluent on EliA Cardiolipin well IgM was around 18 relative fluorescence units (RU). The carry over could hardly be expressed in U/mL since the lowest calibration point 0 U/mL is normally found around 19 RU.

A difference between instrument dilution 1:2 and manual dilution 1:2 could hardly be detected. Therefore after pipetting the sample with the high titer of specific anti-Cardiolipin antibodies, only a few RUs difference compared to the prediluted sample pipetting could be seen, which is too low to be expressed in U/mL.

## g. Assay cut-off:

A study was done on 400 apparently healthy Blood Donor samples from Caucasian individuals equally distributed by sex and age. The 95th percentile should be calculated and should be taken into account for setting of the cut-off. The results are summarized below:

{11}

Cardiolipin IgG/IgM:

|  Test | Unit | No. of samples | Mean value | 95%- percentile | 99%- percentile  |
| --- | --- | --- | --- | --- | --- |
|  EliA Cardiolipin IgG | GPL-U/ml | 400 | 4.6 | 5.7 | 23.8  |

Cut-off:

|  < 10 GPL-U/mL | Negative  |
| --- | --- |
|  10 – 40 GPL-U/mL | Weak positive  |
|  > 40 GPL-U/mL | Positive  |
|  Test | Unit | No. of samples | Mean value | 95%- percentile | 99%- percentile  |
| --- | --- | --- | --- | --- | --- |
|  EliA Cardiolipin IgM | MPL-U/ml | 400 | 4.1 | 11.7 | 29.9  |

Cut-off:

|  < 10 MPL-U/mL | Negative  |
| --- | --- |
|  10 – 40 MPL-U/mL | Weak positive  |
|  > 40 MPL-U/mL | Positive  |

β2-Glycoprotein I IgG/IgM:

The expected value in the normal population is negative. However, up to 3% of apparently healthy, asymptomatic individuals may have increased levels of β2-glycoprotein I antibodies. Expected values may vary depending on the population tested.

|  Test | Unit | No. of samples | Mean value | 95%- percentile | 99%- percentile  |
| --- | --- | --- | --- | --- | --- |
|  EliA β2-Glycoprotein I IgG | U/ml | 400 | 3.3 | 8.2 | 17.7  |

Cut-off:

|  < 7 U/mL | Negative  |
| --- | --- |
|  7 – 10 U/mL | Equivocal  |
|  > 10 U/mL | Positive  |
|  Test | Unit | No. of samples | Mean value | 95%- percentile | 99%- percentile  |
| --- | --- | --- | --- | --- | --- |
|  EliA β2-Glycoprotein I IgM | U/ml | 400 | 1.0 | 3.2 | 5.7  |

Cut-off:

|  < 7 U/mL | Negative  |
| --- | --- |
|  7 – 10 U/mL | Equivocal  |
|  > 10 U/mL | Positive  |

{12}

2. Comparison studies:

a. Method comparison with predicate device:

289 serum samples were collected from the serum bank at Phadia GmbH. In this study 64 samples from patients who had been clinically defined as suffering from Antiphospholipid Syndrome (APS) were included. Each sample was analyzed in duplicates in EliA Cardiolipin IgG kits. The study was evaluated by excluding the clinical samples below the limit of detection and above the measuring range. All method comparison studies were run on ImmunoCAP 250 instruments. The results are summarized below:

EliA Cardiolipin tests, in contrast to the Varelisa Cardiolipin tests, do not yield equivocal, but weak positive results. When evaluating the method comparison the area between 10 and 40 GPL- / MPL-U/ml were considered only as positive (&gt; 10 GPL-U/mL or MPL-U/mL).

EliA Cardiolipin IgG Well - Equivocal Varelisa results considered as negative:

|  n=275 | Varelisa Cardiolipin IgG Abs.  |   |   |   |
| --- | --- | --- | --- | --- |
|   |   |  >15 GPL-U/mL | <15 GPL-U/mL | Total  |
|  EliA Cardiolipin IgG | >10 GPL-U/mL | 37 | 13 | 50  |
|   |  <10 GPL-U/mL | 6 | 219 | 225  |
|   |  Total | 43 | 232 | 275  |

Positive Percent Agreement 86% (37/43) (95% CI: 72.1% - 94.7%)
Negative Percent Agreement 94.4% (219/232) (95%CI: 90.6% - 97.0%)
Total Agreement 93.1% [(37+219)/275] (95%CI: 89.4% - 95.8%)

EliA Cardiolipin IgG Well - Equivocal Varelisa results considered as positive

|  n=275 | Varelisa Cardiolipin IgG Abs.  |   |   |   |
| --- | --- | --- | --- | --- |
|   |   |  >10 GPL-U/mL | <10 GPL-U/mL | Total  |
|  EliA Cardiolipin IgG | >10 GPL-U/mL | 41 | 9 | 50  |
|   |  <10 GPL-U/mL | 20 | 205 | 225  |
|   |  Total | 61 | 214 | 275  |

Positive Percent Agreement 67.2% (41/61) (95% CI: 54.0% - 78.7%)
Negative Percent Agreement 95.8% (205/214) (95%CI: 92.2% - 98.1%)
Total Agreement 89.5% [(41+205)/275] (95%CI: 85.2% - 92.8%)

13

{13}

EliA Cardiolipin IgM Well: Equivocal Varelisa results considered as negative:

|  n=230 | Varelisa Cardiolipin IgM Abs.  |   |   |   |
| --- | --- | --- | --- | --- |
|   |   |  >15 MPL-U/mL | <15 MPL-U/mL | Total  |
|  EliA Cardiolipin IgM | >10 MPL-U/mL | 17 | 25 | 42  |
|   |  <10 MPL-U/mL | 3 | 185 | 188  |
|   |  Total | 20 | 210 | 230  |

Positive Percent Agreement 85% (17/20) (95% CI: 62.1% – 96.8%)
Negative Percent Agreement 88.1% (185/210) (95%CI: 82.9% – 92.1%)
Total Agreement 87.8% [(17+185)/230] (95%CI: 82.9% – 91.8%)

EliA Cardiolipin IgM Well with equivocal Varelisa results considered as positive:

|  n=230 | Varelisa Cardiolipin IgM Abs.  |   |   |   |
| --- | --- | --- | --- | --- |
|   |   |  >10 MPL-U/mL | <10 MPL-U/mL | Total  |
|  EliA Cardiolipin IgM | >10 MPL-U/mL | 23 | 19 | 42  |
|   |  <10 MPL-U/mL | 8 | 180 | 188  |
|   |  Total | 31 | 199 | 230  |

Positive Percent Agreement 74.2% (23/31) (95% CI: 55.4% – 88.1%)
Negative Percent Agreement 90.5% (180/199) (95%CI: 85.5% – 94.2%)
Total Agreement 88.3% [(23+180)/230] (95%CI: 83.4% – 92.1%)

EliA β2-Glycoprotein I IgG

Equivocals EliA 7-10 U/mL - 7.7% Varelisa 7-15 U/mL - 12.1%

EliA β2-Glycoprotein I IgG Well: equivocal samples considered as negative:

|  n=297 | Varelisa β2-GP I IgG Abs.  |   |   |   |
| --- | --- | --- | --- | --- |
|   |   |  >15 MPL-U/mL | <15 MPL-U/mL | Total  |
|  EliA β2-GP I IgG | >10 MPL-U/mL | 36 | 22 | 58  |
|   |  <10 MPL-U/mL | 7 | 232 | 239  |
|   |  Total | 43 | 254 | 297  |

Positive Percent Agreement 83.7% (36/43) (95% CI: 69.3% – 93.2%)
Negative Percent Agreement 91.3% (232/254) (95%CI: 87.2% – 94.5%)
Total Agreement 90.2% [(36+232)/297] (95%CI: 86.3% – 93.4%)

14

{14}

EliA β2-Glycoprotein I IgG Well with equivocal samples considered as positive:

|  n=297 | Varelisa β2-GP I IgG Abs.  |   |   |   |
| --- | --- | --- | --- | --- |
|   |   |  >10 MPL-U/mL | <10 MPL-U/mL | Total  |
|  EliA β2-GP I IgG | >10 MPL-U/mL | 61 | 20 | 81  |
|   |  <10 MPL-U/mL | 10 | 206 | 216  |
|   |  Total | 71 | 226 | 297  |

Positive Percent Agreement 85.9% (61/71) (95% CI: 75.6% - 93.0%)
Negative Percent Agreement 91.2% (206/226) (95%CI: 86.7% - 94.5%)
Total Agreement 89.9% [(61+206)/297] (95%CI: 85.9% - 93.1%)

EliA β2-Glycoprotein I IgM

Equivocals

EliA 7-10 U/mL - 9.4%

Varelisa 7-15 U/mL 21.9%

EliA β2-Glycoprotein I IgM Well with equivocal samples considered as negative:

|  n=128 | Varelisa β2-GP I IgM Abs.  |   |   |   |
| --- | --- | --- | --- | --- |
|   |   |  >15 MPL-U/mL | <15 MPL-U/mL | Total  |
|  EliA β2-GP I IgM | >10 MPL-U/mL | 24 | 19 | 43  |
|   |  <10 MPL-U/mL | 0 | 85 | 85  |
|   |  Total | 24 | 104 | 128  |

Positive Percent Agreement 100% (24/24) (95% CI: 85.8% - 100%)
Negative Percent Agreement 81.7% (85/104) (95%CI: 72.9% - 88.6%)
Total Agreement 85.2% [(24+85)/128] (95%CI: 77.8% - 90.8%)

EliA β2-Glycoprotein I IgM Well with equivocal samples considered as positive:

|  n=128 | Varelisa β2-GP I IgM Abs.  |   |   |   |
| --- | --- | --- | --- | --- |
|   |   |  >10 MPL-U/mL | <10 MPL-U/mL | Total  |
|  EliA β2-GP I IgM | >10 MPL-U/mL | 40 | 15 | 55  |
|   |  <10 MPL-U/mL | 1 | 72 | 73  |
|   |  Total | 41 | 87 | 128  |

Positive Percent Agreement 97.6% (40/41) (95% CI: 87.1% - 99.9%)
Negative Percent Agreement 82.8% (72/87) (95%CI: 73.2% - 90.0%)
Total Agreement 87.5% [(40+72)/128] (95%CI: 80.5% - 92.7%)

{15}

# Summary:

Equivocal results were considered as negative.

|  Product | Predicate Device | Number of samples analyzed | Total agreement [%] | 95% CI  |
| --- | --- | --- | --- | --- |
|  EliA Cardiolipin IgG | Varelisa Cardiolipin IgG Abs. | 275 | 93.1 | 89.4 – 95.8  |
|  EliA Cardiolipin IgM | Varelisa Cardiolipin IgM Abs. | 230 | 87.8 | 82.9 – 91.8  |
|  EliA β2-Glycoprotein I IgG | Varelisa β2-GP I IgG Abs. | 297 | 90.2 | 86.3 – 93.4  |
|  EliA β2-Glycoprotein I IgM | Varelisa β2-GP I IgM Abs. | 128 | 85.2 | 77.8 – 90.8  |

Equivocal results were considered as positive.

|  Product | Predicate Device | Number of samples analyzed | Total agreement [%] | 95% CI  |
| --- | --- | --- | --- | --- |
|  EliA Cardiolipin IgG | Varelisa Cardiolipin IgG Abs. | 275 | 89.5 | 85.2 – 92.8  |
|  EliA Cardiolipin IgM | Varelisa Cardiolipin IgM Abs. | 230 | 88.3 | 83.4 – 92.1  |
|  EliA β2-Glycoprotein I IgG | Varelisa β2-GP I IgG Abs. | 297 | 89.9 | 85.9 – 93.1  |
|  EliA β2-Glycoprotein I IgM | Varelisa β2-GP I IgM Abs. | 128 | 87.5 | 80.5 – 92.7  |

b. Instrument comparison:

A study was done to demonstrate that the performance of  $\mathrm{EliA}^{\mathrm{TM}}$  Cardiolipin IgG,  $\mathrm{EliA}^{\mathrm{TM}}$  Cardiolipin IgM,  $\mathrm{EliA}^{\mathrm{TM}}$ $\beta 2$ -Glycoprotein I IgG and  $\mathrm{EliA}^{\mathrm{TM}}$ $\beta 2$ -Glycoprotein I IgM is equivalent on the Instrument Phadia 100 and Phadia 250.

The results for 36 samples (4 negative samples and 32 positive samples) distributed over the measuring range were determined. All samples were run on three different instruments in two runs and single replicates (6 independent values for Phadia 250, 6 independent values for Phadia 100). Results are summarized below:

|   | Slope (95% CI) | Y-intercept(95% CI)  |
| --- | --- | --- |
|  EliA Cardiolipin IgG | 1.016 (0.969 to 1.67) | -0.984 (-2.464 to 0.194)  |
|  EliA Cardiolipin IgM | 0.901 (0.877 to 0.928) | -0.607 (-1.023 to -0.217)  |
|  EliA β2-Glycoprotein I IgG | 1.008 (0.963 to 1.093) | -0.626 (-4.188 to 1.539)  |
|  EliA β2-Glycoprotein I IgM | 0.996 (0.964 to 1.038) | 0.425 (-1.435 to 0.368)  |

b. Matrix comparison: The purpose of this study was to demonstrate that the new devices give the same results for Serum, Heparin plasma, Citrate plasma and EDTA plasma collected from the same patient. The study was done on 50 samples, 25 negative and 25 positive, spread across the assay range. Samples were run in single replicates. All the method comparison studies were run on ImmunoCAP 250 instruments. The results are summarized below:

{16}

|  test | No of. Samples | Range tested | Slope | Y.intercept | R2  |
| --- | --- | --- | --- | --- | --- |
|  EliA Cardiolipin IgG |   | GPL-U/ml | 95% CI | 95% CI |   |
|  Serum v plasma citrate | 24 pos/ 26 neg., 8 low pos* | 1 - 522 | 0,99 (0,98 to 1,01) | 0,93 (-1,12 to 2,98) | 1  |
|  Serum v plasma EDTA | 24 pos/ 26 neg., 8 low pos* | 1,1 - 484 | 1,01 (0,99 to 1,02) | -0,43 (-2,04 to 1,20) | 1  |
|  Serum v plasma heparin | 24 pos/ 26 neg., 8 low pos* | 1,4 - 490 | 1,00 (0,98 to 1,02) | -0,74 (-3,09 to 1,60) | 1  |
|  EliA Cardiolipin IgM |   | MPL-U/ml | 95% CI | 95% CI |   |
|  Serum v plasma citrate | 24 pos/ 26 neg., 8 low pos* | 1 - 522 | 0,99 (0,98 to 1,01) | 0,93 (-1,12 to 2,98) | 1  |
|  Serum v plasma EDTA | 24 pos/ 26 neg., 8 low pos* | 1,1 - 484 | 1,01 (0,99 to 1,02) | -0,43 (-2,04 to 1,20) | 1  |
|  Serum v plasma heparin | 24 pos/ 26 neg., 8 low pos* | 1,4 - 490 | 1,00 (0,98 to 1,02) | -0,74 (-3,09 to 1,60) | 1  |
|  EliA β2-GPI IgG |   | U/ml | 95% CI | 95% CI |   |
|  Serum v plasma citrate | 24 pos/ 26 neg., 0 equiv. | 0,1 - 488 | 0,93 (0,91 to 0,95) | 1,02 (-2,28 to 4,31) | 0.99  |
|  Serum v plasma EDTA | 25 pos/ 26 neg., 0 equiv. | 0,4 - 441 | 1,04 (1,01 to 1,07) | -0,69 (-4,72 to 3,35) | 0.99  |
|  Serum v plasma heparin | 26 pos/ 26 neg., 0 equiv. | 0,5 - 525 | 0,90 (0,88 to 0,91) | 3,5 (0,85 to 6,14) | 1  |
|  EliA β2-GPI IgM |   | U/ml | 95% CI | 95% CI |   |
|  Serum v plasma citrate | 24 pos/ 26 neg., 0 equiv. | 0,0 - 469 | 0,90 (0,89 to 0,92) | 1,9 (-0,46 to 4,3) | 1  |
|  Serum v plasma EDTA | 25 pos/ 26 neg., 0 equiv. | 0,0 - 446 | 1,00 (0,99 to 1,02) | 0,14 (-2,41 to 2,69) | 1  |
|  Serum v plasma heparin | 26 pos/ 26 neg., 0 equiv. | 0,0 - 463 | 0,92 (0,91 to 0,94) | -148 (-3,79 to 0,83) | 1  |

{17}

18

3. Clinical studies:

a. Clinical Sensitivity and Specificity:

The samples that were used for the method comparison were used to calculate the sensitivity/specificity of the assay. SLE samples with unknown APS status were also evaluated for their sensitivity/specificity.

Clinical Evaluation of EliA Cardiolipin IgG

|  EliA Cardiolipin IgG | Diagnostic Group APS  |   |   |
| --- | --- | --- | --- |
|  n=224 | + | - | Total  |
|  Positive test > 10 GPL U/mL | 23 | 4 | 27  |
|  Negative test ≤ 10 GPL U/mL | 26 | 171 | 197  |
|  Total | 49 | 175 | 224  |
|  Sensitivity: | 46.9% | 95% CI: 32.5 – 61.7 %  |
| --- | --- | --- |
|  Specificity: | 97.7% | 95% CI: 94.3 – 99.4 %  |

Clinical Evaluation of EliA Cardiolipin IgM

|  EliA Cardiolipin IgM | Diagnostic Group APS  |   |   |
| --- | --- | --- | --- |
|  n=188 | + | - | Total  |
|  Positive test > 10 MPL U/ml | 14 | 10 | 24  |
|  Negative test ≤ 10 MPL U/ml | 45 | 119 | 164  |
|  Total | 59 | 129 | 188  |
|  Sensitivity: | 23.7% | 95% CI: 13.6 – 36.6 %  |
| --- | --- | --- |
|  Specificity: | 92.2% | 95% CI: 86.2 – 96.2 %  |

Clinical Evaluation of EliA β2-Glycoprotein I IgG

|  EliA β2-GPI IgG | Diagnostic Group APS  |   |   |
| --- | --- | --- | --- |
|  n=220 | + | - | Total  |
|  Positive test > 10 U/ml | 24 | 2 | 26  |
|  Negative test ≤ 10 U/ml | 30 | 164 | 194  |
|  Total | 54 | 166 | 220  |
|  Sensitivity: | 44.4% | 95% CI: 30.9 – 58.6 %  |
| --- | --- | --- |
|  Specificity: | 98.8% | 95% CI: 95.7 – 99.9 %  |

{18}

Clinical Evaluation of EliA β2-Glycoprotein I IgM

|  EliA β2-GPI IgM | Diagnostic Group APS  |   |   |
| --- | --- | --- | --- |
|  n=77 | + | - | Total  |
|  Positive test >10 U/ml | 8 | 4 | 12  |
|  Negative test ≤10 U/ml | 25 | 40 | 65  |
|  Total | 33 | 44 | 77  |
|  Sensitivity: | 24.2% | 95% CI: 11.1 – 42.3 %  |
| --- | --- | --- |
|  Specificity: | 90.9% | 95% CI: 78.3 – 97.5 %  |

The table below shows the results for each clinical subgroup:

|  Condition | β-2 Glycoprotein I IgG % positive (>10U/mL) | β-2 Glycoprotein I IgG % positive (>10U/mL) | Cardiolipin IgG % positive (>10U/mL) | Cardiolipin IgM % positive (>10U/mL)  |
| --- | --- | --- | --- | --- |
|  |   |   |   |   |
|  Anti-Phospholipid Syndrome (APS) (37 samples from primary APS) | 34/64=53% | 8/64=12.5% | 23/49=46.9% | 14/60=23.3%  |
|  Systemic lupus Erythematosus (SLE) | 7/41=17.0% | 9/41=22.0% | 10/37=27.0% | 5/29=17.2%  |
|  Unknown | 27/37=73.0% | 27/37=73.0% | 13/14=92.9% | 13/13=100%  |
|  1Infectious diseases | 1/112=0.89% | 2/112=1.8% | 2/11=11.8% | 7/84=8.3%  |
|  2Other | 1/72=1.4% | 2/72=2.8% | 2/64=3.1% | 3/47=6.4%  |

$^{1}$ Infectious diseases include: Hepatitis C Virus (HCV), HIV, Epstein Barr Virus (EBV), Helicobacter pylori infection (HBP), Syphilis, Toxoplasmosis (TP), Varicella Zoster Virus (VZV), Yersinia Pestis (YP)

$^{2}$ Other include: Marine-Lenhart Syndrome (MLS), Secondary Sjögren's Syndrome (SSS), Chronic lymphocytic Thyroiditis, atropic (clTHy), Hashimoto's disease (HM), Hypothyreosis (Hypthy), Morbus Basedow (MB), Morbus Basedow endocrine ophthalmopathy (MBeO), Mixed Connective Tissue Disease (MCTD), Primary Sjögren's Syndrome (PSS), Rheumatoid Arthritis (RA), Rheum factor (RF), CM

b. Other clinical supportive data (when a. is not applicable):

4. Clinical cut-off:

See assay cut-off

5. Expected values/Reference range: : Cardiolipin IgG/IgM:

{19}

The expected value in the normal population is negative. The sponsor states that a low percentage (up to 5 %) of apparently healthy, asymptomatic individuals have been reported to be positive for Anti-Cardiolipin Antibody. Their proportion may increase with age. Expected values may vary depending on the population tested.

**β2-Glycoprotein I IgG/IgM:**
The expected value in the normal population is negative. The sponsor states that up to 3% of apparently healthy, asymptomatic individuals may have increased levels of β2-glycoprotein I antibodies. Expected values may vary depending on the population tested.

**N. Proposed Labeling:**
The labeling is sufficient and it satisfies the requirements of 21 CFR Part 809.10.

**O. Conclusion: SE**
The submitted information in this premarket notification is complete and supports a substantial equivalence decision.

20

---

**Source:** [https://fda.innolitics.com/submissions/IM/subpart-f%E2%80%94immunological-test-systems/MID/K091845](https://fda.innolitics.com/submissions/IM/subpart-f%E2%80%94immunological-test-systems/MID/K091845)

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