Predicate Devices

Reference Devices

Not Found

AI/ML (Artificial Intelligence / Machine Learning)

No
The summary describes a standard automated immunoassay system that uses predefined master and working curves for calculation, without mentioning any AI or ML components.

Therapeutic

No
This device is an in vitro diagnostic (IVD) immunoassay designed for the semi-quantitative determination of antibodies to aid in the diagnosis of Systemic Lupus Erythematosus (SLE) and Mixed Connective Tissue Disease (MCTD). It is used for diagnostic purposes, not for treating diseases, and therefore is not a therapeutic device.

Diagnostic

Yes

The "Intended Use / Indications for Use" section explicitly states that the device's outputs "can aid in the diagnosis of Systemic Lupus Erythematosus (SLE)" and "Mixed Connective Tissue Disease (MCTD)."

SaMD (Software as a Medical Device)

No

The device description explicitly states that the system includes "liquid handling hardware, luminometer and computer with software-user interface" and utilizes "reagent cartridge format". This indicates the device is a physical instrument with associated reagents, not solely software.

IVD (In Vitro Diagnostic)

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

Here's why:

Intended Use: The intended use explicitly states the device is for the "semi-quantitative determination of IgG anti-Sm antibodies in human serum" and "IgG anti-ribonucleoprotein (RNP) antibodies in human serum." It also states that the presence of these antibodies, in conjunction with clinical findings and other laboratory tests, can "aid in the diagnosis of Systemic Lupus Erythematosus (SLE)" and "Mixed Connective Tissue Disease (MCTD)." This clearly indicates the device is intended for use in the diagnosis of disease.
Sample Type: The device uses "human serum" as the sample type.
Method: The device employs a "chemiluminescent immunoassay," which is a laboratory test performed in vitro (outside the body).
Device Description: The description details the reagents and process for performing the assay on the BIO-FLASH instrument, which is a laboratory instrument.
Components: The kit includes reagents, calibrators, and controls, which are typical components of IVD kits used in laboratory testing.
Performance Studies: The document includes detailed performance studies such as precision, reproducibility, limit of detection, analytical measuring range, interference, cross-reactivity, stability, and clinical sensitivity and specificity. These are standard evaluations for IVD devices.
Comparison with Predicate Device: The comparison to a predicate device (QUANTA Lite™ Sm ELISA) further confirms its nature as a diagnostic test.

All of these factors align with the definition of an In Vitro Diagnostic device, which is a medical device intended for use in vitro for the examination of specimens, including blood and tissue samples, derived from the human body, solely or principally for the purpose of providing information concerning a physiological or pathological state, or concerning a congenital abnormality, or to determine the compatibility of transplant tissues, organs or blood, or to monitor therapeutic measures.

PCCP (Predetermined Change Control Plan) Authorized

N/A

Overview

Intended Use / Indications for Use

The QUANTA Flash Sm is a chemiluminescent immunoassay for the semi-quantitative determination of lgG anti-Sm antibodies in human serum. The presence of antibodies, in conjunction with clinical findings and other laboratory tests, can aid in the diagnosis of Systemic Lupus Erythematosus (SLE).

The QUANTA Flash RNP is a chemiluminescent immunoassay for the semi-quantitative determination of lgG anti-ribonucleoprotein (RNP) antibodies in human serum. The presence of anti-RNP antibodies, in conjunction with clinical findings and other laboratory tests, can aid in the diagnosis of Systemic Lupus Erythematosus (SLE) and Mixed Connective Tissue Disease (MCTD).

QUANTA Flash Sm Calibrators are intended for use with the QUANTA Flash Sm chemiluminescent immunoassay for the determination of IgG anti-Sm antibodies in human serum. Each calibrator establishes a point of reference for the working curve that is used to calculate unit values.

QUANTA Flash RNP Calibrators are intended for use with the QUANTA Flash RNP chemiluminescent immunoassay for the determination of IgG anti-RNP antibodies in human serum. Each calibrator establishes a point of reference for the working curve that is used to calculate unit values.

QUANTA Flash Sm Controls are intended for use with the QUANTA Flash Sm chemiluminescent immunoassay for quality control in the determination of IgG anti-Sm antibodies in human serum.

QUANTA Flash RNP Controls are intended for use with the QUANTA Flash RNP chemiluminescent immunoassay for quality control in the determination of IgG anti-RNP antibodies in human serum.

Product codes (comma separated list FDA assigned to the subject device)

LKP, LKO, JIT, JJX

Device Description

The QUANTA Flash Sm and RNP assays are designed to run on the BIO-FLASH instrument. This platform is a fully automated closed system with continuous load and random access capabilities that automatically processes the samples, runs the assay and reports the results. It includes liquid handling hardware, luminometer and computer with software-user interface. The QUANTA Flash Sm and RNP assays utilize a reagent cartridge format, which is compatible with the BIO-FLASH instrument.

Native Sm or RNP antigen that is purified from calf thymus is coated onto paramagnetic beads. The bead suspension is lyophilized and stored in the bead tube. Prior to use in the BIO-FLASH system, the sealed reagent tubes are pierced with the reagent cartridge lid and the beads are rehydrated and resuspended using resuspension buffer by pipetting up and down with a transfer pipette. The reagent cartridge is then loaded onto the BIO-FLASH instrument. Samples are also loaded onto the instrument in sample racks. A patient serum sample is prediluted by the BIO-FLASH with system rinse in a small disposable plastic cuvette. Small amounts of the diluted patient serum, the beads, and assay buffer are all combined into a second cuvette, and mixed. This cuvette is then incubated at 37°C. The beads are magnetized and washed several times. Isoluminol conjugated anti-human IgG antibodies are then added to the cuvette, and again incubated at 37°C. The beads are magnetized and washed repeatedly. The isoluminol conjugate is oxidized when Trigger 1 (Fe(III)coproporphyrin in sodium hydroxide solution) and Trigger 2 (urea-hydrogen peroxide in sodium chloride solution) are added to the cuvette, and the flash of light produced from this reaction is measured as Relative Light Units (RU) by the BIO-FLASH optical system. The RLU are proportional to the amount of isoluminol conjugate that is bound to the human lgG, which is in turn proportional to the amount of anti-RNP antibodies bound to the Sm or RNP on the beads.

For quantitation, the QUANTA Flash Sm and RNP assays utilize a predefined lot specific Master Curve that is uploaded onto the instrument through the reagent cartridge barcode. Every new lot number of reagent cartridge must be calibrated before first use with the QUANTA Flash Sm and RNP Calibrators. Based on the results obtained with the two Calibrators included in the Calibrator Set (sold separately), an instrument specific Working Curve is created, which is used to calculate chemiluminescent units (CU) from the instrument signal (RLU) obtained for each sample.

The QUANTA Flash Sm kit contains the following materials:
One (1) QUANTA Flash Sm Reagent Cartridge
One (1) vial of Resuspension buffer
One (1) Transfer pipette

The QUANTA Flash Sm reagent cartridge contains the following reagents for 50 determinations:

Sm antigen coated paramagnetic beads, lyophilized. a.
Assay buffer colored pink, containing Tris-buffered saline, Tween 20, protein b. stabilizers and preservatives.
Tracer IgG Isoluminol labeled anti-human IgG antibodies in buffer, containing protein C. stabilizers and preservative.

The QUANTA Flash RNP kit contains the following materials:
One (1) QUANTA Flash RNP Reagent Cartridge

One (1) vial of Resuspension buffer
One (1) Transfer pipette

The QUANTA Flash RNP reagent cartridge contains the following reagents for 50 determinations:

RNP antigen coated paramagnetic beads, lyophilized. a.
Assay buffer colored pink, containing Tris-buffered saline, Tween 20, protein b. stabilizers and preservatives.
Tracer IgG Isoluminol labeled anti-human IgG antibodies in buffer, containing protein C. stabilizers and preservative.

The QUANTA Flash Sm Calibrators kit and the QUANTA Flash™ RNP Calibrators kit each contain 2 vials of Calibrators:

QUANTA Flash Sm Calibrators:

QUANTA Flash Sm Calibrator 1: Two (2) barcode labeled tubes containing 0.3 ml prediluted, ready to use reagent. Calibrators contain human antibodies to Sm in buffer, protein stabilizers, and preservatives.
QUANTA Flash Sm Calibrator 2: Two (2) barcode labeled tubes containing 0.3 mL prediluted, ready to use reagent. Calibrators contain human antibodies to Sm in buffer, protein stabilizers, and preservatives.

QUANTA Flash RNP Calibrators:

QUANTA Flash RNP Calibrator 1: Two (2) barcode labeled tubes containing 0.3 mL prediluted, ready to use reagent. Calibrators contain human antibodies to RNP in buffer,
protein stabilizers, and preservatives.
QUANTA Flash RNP Calibrator 2: Two (2) barcode labeled tubes containing 0.3 mL prediluted, ready to use reagent. Calibrators contain human antibodies to RNP in buffer, protein stabilizers, and preservatives.

The QUANTA Flash Sm Controls kit and the QUANTA Flash™ RNP Controls kit each contain 2 vials of Negative Control and two vials of Positive Control:

QUANTA Flash Sm Controls:

QUANTA Flash™ Sm Negative Control: Two (2) barcode labeled tubes containing 0.5 mL, ready to use reagent. Controls contain human antibodies to Sm in buffer, protein stabilizers, and preservatives.
QUANTA Flash™ Sm Positive Control: Two (2) barcode labeled tubes containing 0.5 ml, ـــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــ ready to use reagent. Controls contain human antibodies to Sm in buffer, protein stabilizers, and preservatives.

QUANTA Flash RNP Controls:

QUANTA Flash™ RNP Negative Control: Two (2) barcode labeled tubes containing 0.5 mL, ready to use reagent. Controls contain human antibodies to RNP in buffer, protein stabilizers, and preservatives.
QUANTA Flash™ RNP Positive Control: Two (2) barcode labeled tubes containing 0.5 ml, ready to use reagent. Controls contain human antibodies to RNP in buffer, protein stabilizers, and preservatives.

Mentions image processing

Not Found

Mentions AI, DNN, or ML

Not Found

Input Imaging Modality

Not Found

Anatomical Site

Human serum

Indicated Patient Age Range

Not Found

Intended User / Care Setting

Not Found

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)

Precision (repeatability)

QUANTA Flash Sm:
- Sample ID 1: N=80, Mean=13.1 CU, Total Precision (SD=1.1, %CV=8.7)
- Sample ID 2: N=80, Mean=22.0 CU, Total Precision (SD=2.5, %CV=11.5)
- Sample ID 3: N=84, Mean=93.0 CU, Total Precision (SD=7.7, %CV=8.3)
- All %CV values were within the acceptance limit, 15%.
QUANTA Flash RNP:
- Sample ID 1: N=92, Mean=6.7 CU, Total Precision (SD=0.4, %CV=6.4)
- Sample ID 2: N=84, Mean=24.8 CU, Total Precision (SD=1.2, %CV=4.8)
- Sample ID 3: N=88, Mean=31.6 CU, Total Precision (SD=3.0, %CV=9.4)
- Sample ID 4: N=92, Mean=120.7 CU, Total Precision (SD=8.0, %CV=6.6)
- Sample ID 5: N=88, Mean=218.6 CU, Total Precision (SD=23.6, %CV=10.8)
- Sample ID 6: N=88, Mean=319.5 CU, Total Precision (SD=21.8, %CV=6.8)
- Sample ID 7: N=96, Mean=409.6 CU, Total Precision (SD=35.5, %CV=8.7)
- All %CV values were within the acceptance limit, 15%.

Reproducibility (between reagent lots, calibrator lots, operators)

QUANTA Flash Sm:
- Sample #1: N=80 replicates, Mean=15.2 CU, Total Precision (SD=1.1, CV=7.0%)
- Sample #2: N=80 replicates, Mean=23.3 CU, Total Precision (SD=1.8, CV=7.7%)
- Sample #3: N=80 replicates, Mean=157.3 CU, Total Precision (SD=13.0, CV=8.3%)
- All %CV values were within the acceptance limit, 15%.
QUANTA Flash RNP:
- Sample #1: N=80 replicates, Mean=15.3 CU, Total Precision (SD=0.4, CV=2.8%)
- Sample #2: N=80 replicates, Mean=23.1 CU, Total Precision (SD=0.6, CV=2.6%)
- Sample #3: N=80 replicates, Mean=173.1 CU, Total Precision (SD=5.1, CV=2.9%)
- All %CV values were within the acceptance limit, 15%.

Limit of Blank (LoB), Limit of Detection (LoD)

QUANTA Flash Sm:
- LoD = 803 RLU
- LoB = 540 RLU
QUANTA Flash RNP:
- LoD = 1128.8 RLU
- LoB = 713 RLU

Analytical Measuring Range (AMR) / Linearity

QUANTA Flash Sm:
- AMR: 3.3 CU to 693.5 CU (linear range)
- Linearity study: 5 serum samples, R2 = 0.99
QUANTA Flash RNP:
- AMR: 3.5 CU to 643.8 CU (linear range)
- Linearity study: 5 serum samples, R2 = 1.00

Auto-Rerun Function Validation

QUANTA Flash Sm:
- 5 high positive specimens tested.
- % recovery values for auto-rerun vs. manual dilution: 87.6% to 97.7% (average 91.5%).
QUANTA Flash RNP:
- 6 high positive specimens tested.
- % recovery values for auto-rerun vs. manual dilution: 82.5% to 102.2% (average 96.3%).
All recoveries within ±20% acceptance limit.

Interference

QUANTA Flash Sm: No interference detected with bilirubin (up to 10 mg/dL), hemoglobin (up to 200 mg/dL), triglycerides (up to 1000 mg/dL), cholesterol (up to 224.3 mg/dL), and RF IgM (up to 500 IU/mL). Recovery within 85%-115% or ±4 CU.
QUANTA Flash RNP: No interference detected with bilirubin (up to 10 mg/dL), hemoglobin (up to 200 mg/dL), triglycerides (up to 1000 mg/dL), cholesterol (up to 224.3 mg/dL), and RF IgM (up to 500 IU/mL). Recovery within 85%-115% or ±4 CU.

High concentration hook effect

No hook effect observed up to 2429 CU for Sm assay and up to 3140 CU for RNP assay.

Cross-reactivity

QUANTA Flash Sm: 233 patient samples tested. 5 positive samples (3 scleroderma, 1 systemic rheumatic disease, 1 infectious disease HIV). Overall 2.1% positive.
QUANTA Flash RNP: 246 patient samples tested. 12 positive samples (8 scleroderma, 2 rheumatoid arthritis, 1 poly- and dermatomyositis, 1 infectious disease HIV). Overall 4.9% positive.

Stability

Shelf life (accelerated studies):
- Sm beads, RNP beads, Sm Calibrators, RNP Calibrators, Sm Controls, RNP Controls met acceptance criteria for 1-year preliminary expiration.
In-use (onboard) stability:
- Sm Calibrators: 4 calibrations over an 8-hour period.
- Sm Controls: 15 uses, up to 2.5 hours total.
- Sm Reagent Cartridge: 33 days.
- RNP Calibrators: 4 calibrations over an 8-hour period.
- RNP Controls: 15 uses, up to 2.5 hours total.
- RNP Reagent Cartridge: 28 days.

Reference Ranges/Cut-off

QUANTA Flash Sm: Cut-off = 20 CU (Negative

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

K123593

APR 1 7 2013

510(k) Summary

This summary of the 510(k) safety and effectiveness information is being submitted in accordance with the requirements of SMDA 1990 and 21 CFR 807.92.

| Submitter: | INOVA Diagnostics, Inc
9900 Old Grove Road,
San Diego, CA, 92131 | |
|----------------------------------|---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|--------------------------------------------------------------------------------------------------------------|
| Purpose of submission: | New device(s) | |
| Devices in the submission: | QUANTA Flash® Sm
QUANTA Flash® Sm Calibrators
QUANTA Flash® Sm Controls
QUANTA Flash® RNP
QUANTA Flash® RNP Calibrators
QUANTA Flash® RNP Controls | |
| Scientific contact: | Gabriella Lakos, Director of Research, Rheumatology
INOVA Diagnostics, Inc
9900 Old Grove Road, San Diego, CA, 92131
Phone: 858-586-9900/393
Fax: 858-863-0025
email: glakos@inovadx.com | |
| Quality Systems contact: | Tana Keivens, Director, Quality Systems
INOVA Diagnostics, Inc
9900 Old Grove Road, San Diego, CA, 92131
Phone: 858-586-9900
Fax: 858-863-0025/351
email: tkeivens@inovadx.com | |
| Preparation date: | 11/19/2012 | |
| Device (1) name (assay kit): | Proprietary name: QUANTA Flash® Sm
Common name: Anti-Sm Chemiluminescent Immunoassay
Classification name: anti-Sm antibody, antigen and control | |
| Regulation Description | Antinuclear antibody immunological test system | |
| Regulation Medical Specialty | Immunology | |
| Review Panel | Immunology | |
| Product Code | LKP | |
| Regulation Number | 866.5100 | |
| Device Class | 2 | |
| | Common name: | Sm Calibrators |
| | Classification name: | Calibrator, secondary |
| Regulation Description | Calibrator | |
| Regulation Medical Specialty | Clinical Chemistry | |
| Product Code | JIT | |
| Regulation Number | 862.1150 | |
| Device Class | 2 | |
| Device (1) name (Controls): | Proprietary name: | QUANTA Flash® Sm Controls |
| | Common name: | Sm Controls |
| | Classification name: | Single (specified) analyte controls (assayed and unassayed) |
| Regulation Description | Quality control material (assayed and unassayed) | |
| Regulation Medical Specialty | Clinical Chemistry | |
| Product Code | JJX | |
| Regulation Number | 862.1660 | |
| Device Class | 1 | |
| Predicate device (for Device 1): | QUANTA Lite™ Sm ELISA, 510(k) number: K922831 | |
| Device (2) name (assay kit): | Proprietary name: | QUANTA Flash® RNP |
| | Common name: | Anti-RNP Chemiluminescent Immunoassay |
| | Classification name: | anti-RNP antibody, antigen and control |
| Regulation Description | Antinuclear antibody immunological test system | |
| Regulation Medical Specialty | Immunology | |
| Review Panel | Immunology | |
| Product Code | LKO | |
| Regulation Number | 866.5100 | |
| Device Class | 2 | |
| Device (2) name (Calibrators): | Proprietary name: | QUANTA Flash® RNP Calibrators |
| | Common name: | RNP Calibrators |
| | Classification name: | Calibrator, secondary |
| Regulation Description | Calibrator | |
| Product Code | JIT | |
| Regulation Number | 862.1150 | |
| Device Class | 2 | |
| Device (2) name (Controls): | Proprietary name:
Common name:
Classification name: | QUANTA Flash® RNP Controls
RNP Controls
single (specified) analyte controls (assayed and
unassayed) |
| Regulation Description | Quality control material (assayed and unassayed) | |
| Regulation Medical Specialty | Clinical Chemistry | |
| Product Code | JJX | |
| Regulation Number | 862.1660 | |
| Device Class | 1 | |

Device (1) name (Calibrators): Proprietary name:

QUANTA Flash® Sm Calibrators

1

and the comments of the comments of the comments of the comments of

.

:

2

Device (1) and (2) description:

The QUANTA Flash Sm and RNP assays are designed to run on the BIO-FLASH instrument. This platform is a fully automated closed system with continuous load and random access capabilities that automatically processes the samples, runs the assay and reports the results. It includes liquid handling hardware, luminometer and computer with software-user interface. The QUANTA Flash Sm and RNP assays utilize a reagent cartridge format, which is compatible with the BIO-FLASH instrument.

Native Sm or RNP antigen that is purified from calf thymus is coated onto paramagnetic beads. The bead suspension is lyophilized and stored in the bead tube. Prior to use in the BIO-FLASH system, the sealed reagent tubes are pierced with the reagent cartridge lid and the beads are rehydrated and resuspended using resuspension buffer by pipetting up and down with a transfer pipette. The reagent cartridge is then loaded onto the BIO-FLASH instrument. Samples are also loaded onto the instrument in sample racks. A patient serum sample is prediluted by the BIO-FLASH with system rinse in a small disposable plastic cuvette. Small amounts of the diluted patient serum, the beads, and assay buffer are all combined into a second cuvette, and mixed. This cuvette is then incubated at 37°C. The beads are magnetized and washed several times. Isoluminol conjugated anti-human IgG antibodies are then added to the cuvette, and again incubated at 37°C. The beads are magnetized and washed repeatedly. The isoluminol conjugate is oxidized when Trigger 1 (Fe(III)coproporphyrin in sodium hydroxide solution) and Trigger 2 (urea-hydrogen peroxide in sodium chloride solution) are added to the cuvette, and the flash of light produced from this reaction is measured as Relative Light Units (RU) by the BIO-FLASH optical system. The RLU are proportional to the amount of isoluminol conjugate that is bound to the human lgG, which is in turn proportional to the amount of anti-RNP antibodies bound to the Sm or RNP on the beads.

For quantitation, the QUANTA Flash Sm and RNP assays utilize a predefined lot specific Master Curve that is uploaded onto the instrument through the reagent cartridge barcode. Every new lot number of

3

reagent cartridge must be calibrated before first use with the QUANTA Flash Sm and RNP Calibrators. Based on the results obtained with the two Calibrators included in the Calibrator Set (sold separately), an instrument specific Working Curve is created, which is used to calculate chemiluminescent units (CU) from the instrument signal (RLU) obtained for each sample.

The QUANTA Flash Sm kit contains the following materials:

One (1) QUANTA Flash Sm Reagent Cartridge

One (1) vial of Resuspension buffer

One (1) Transfer pipette

The QUANTA Flash Sm reagent cartridge contains the following reagents for 50 determinations:

Sm antigen coated paramagnetic beads, lyophilized. a.
Assay buffer colored pink, containing Tris-buffered saline, Tween 20, protein b. stabilizers and preservatives.
Tracer IgG Isoluminol labeled anti-human IgG antibodies in buffer, containing protein C. stabilizers and preservative.

The QUANTA Flash RNP kit contains the following materials:

One (1) QUANTA Flash RNP Reagent Cartridge

One (1) vial of Resuspension buffer
One (1) Transfer pipette

The QUANTA Flash RNP reagent cartridge contains the following reagents for 50 determinations:

RNP antigen coated paramagnetic beads, lyophilized. a.
Assay buffer colored pink, containing Tris-buffered saline, Tween 20, protein b. stabilizers and preservatives.
Tracer IgG Isoluminol labeled anti-human IgG antibodies in buffer, containing protein C. stabilizers and preservative.

The QUANTA Flash Sm Calibrators kit and the QUANTA Flash™ RNP Calibrators kit each contain 2 vials of Calibrators:

QUANTA Flash Sm Calibrators:

QUANTA Flash Sm Calibrator 1: Two (2) barcode labeled tubes containing 0.3 ml prediluted, ready to use reagent. Calibrators contain human antibodies to Sm in buffer, protein stabilizers, and preservatives.
QUANTA Flash Sm Calibrator 2: Two (2) barcode labeled tubes containing 0.3 mL prediluted, ready to use reagent. Calibrators contain human antibodies to Sm in buffer, protein stabilizers, and preservatives.

QUANTA Flash RNP Calibrators:

QUANTA Flash RNP Calibrator 1: Two (2) barcode labeled tubes containing 0.3 mL prediluted, ready to use reagent. Calibrators contain human antibodies to RNP in buffer,

4

protein stabilizers, and preservatives.

QUANTA Flash RNP Calibrator 2: Two (2) barcode labeled tubes containing 0.3 mL prediluted, ready to use reagent. Calibrators contain human antibodies to RNP in buffer, protein stabilizers, and preservatives.
The QUANTA Flash Sm Controls kit and the QUANTA Flash™ RNP Controls kit each contain 2 vials of Negative Control and two vials of Positive Control:

QUANTA Flash Sm Controls:

QUANTA Flash™ Sm Negative Control: Two (2) barcode labeled tubes containing 0.5 mL, ready to use reagent. Controls contain human antibodies to Sm in buffer, protein stabilizers, and preservatives.
QUANTA Flash™ Sm Positive Control: Two (2) barcode labeled tubes containing 0.5 ml, ـــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــ ready to use reagent. Controls contain human antibodies to Sm in buffer, protein stabilizers, and preservatives.

QUANTA Flash RNP Controls:

QUANTA Flash™ RNP Negative Control: Two (2) barcode labeled tubes containing 0.5 mL, ready to use reagent. Controls contain human antibodies to RNP in buffer, protein stabilizers, and preservatives.
QUANTA Flash™ RNP Positive Control: Two (2) barcode labeled tubes containing 0.5 ml, ready to use reagent. Controls contain human antibodies to RNP in buffer, protein stabilizers, and preservatives.

Intended use(s):

The QUANTA Flash Sm is a chemiluminescent immunoassay for the semi-quantitative determination of lgG anti-Sm antibodies in human serum. The presence of antibodies, in conjunction with clinical findings and other laboratory tests, can aid in the diagnosis of Systemic Lupus Erythematosus (SLE).

The QUANTA Flash RNP is a chemiluminescent immunoassay for the semi-quantitative determination of lgG anti-ribonucleoprotein (RNP) antibodies in human serum. The presence of anti-RNP antibodies, in conjunction with clinical findings and other laboratory tests, can aid in the diagnosis of Systemic Lupus Erythematosus (SLE) and Mixed Connective Tissue Disease (MCTD).

QUANTA Flash Sm Calibrators are intended for use with the QUANTA Flash Sm chemiluminescent immunoassay for the determination of IgG anti-Sm antibodies in human serum. Each calibrator establishes a point of reference for the working curve that is used to calculate unit values.

QUANTA Flash RNP Calibrators are intended for use with the QUANTA Flash RNP chemiluminescent immunoassay for the determination of IgG anti-RNP antibodies in human serum. Each calibrator establishes a point of reference for the working curve that is used to calculate unit values.

QUANTA Flash Sm Controls are intended for use with the QUANTA Flash Sm chemiluminescent immunoassay for quality control in the determination of IgG anti-Sm antibodies in human serum.

QUANTA Flash RNP Controls are intended for use with the QUANTA Flash RNP chemiluminescent immunoassay for quality control in the determination of IgG anti-RNP antibodies in human serum.

5

Substantial equivalence:

The QUANTA Flash Sm, the QUANTA Flash Sm Calibrators and the QUANTA Flash Sm Controls have the same intended use and assay principle as the predicate device.

The QUANTA Flash RNP, the QUANTA Flash RNP Calibrators and the QUANTA Flash RNP Controls have the same intended use and assay principle as the predicate device.

Comparison to predicate device:

Similarities
Item	QUANTA Flash Sm	Predicate Device
Intended use	Semi-quantitative determination of
anti-Sm antibodies in human serum	Semi-quantitative detection of anti-Sm
antibodies in human serum
Assay methodology	Solid phase (heterogenous)
immunoassay	Solid phase (heterogeneous)
immunoassay
Traceability	International Reference Preparation is
not available
Results are traceable to in-house
Standards	International Reference Preparation is
not available
Antigen	Native Sm antigen, purified from calf
thymus	Native Sm antigen, purified from calf
thymus
Sample type	Serum	Serum
Shelf life	One year	One year

QUANTA Flash Sm reagent kit

Differences
Item	QUANTA Flash Sm	Predicate Device
Detection/
Operating principle	Chemiluminescent immunoassay	Enzyme-linked immunosorbent assay
Solid phase	Paramagnetic microparticles (beads)	96-well plate
Conjugate	Isoluminol conjugated anti-human IgG	HRP conjugated anti-human IgG
Calibration	Lot specific Master Curve + two
Calibrators (Sold separately)	Sm ELISA Low Positive
(Included in the kit)

QUANTA Flash RNP reagent kit

Similarities
Item	QUANTA Flash RNP	Predicate Device
Intended use	Semi-quantitative determination of
anti-RNP antibodies in human serum	Semi-quantitative detection of anti-RNP
antibodies in human serum
Assay methodology	Solid phase (heterogenous)
immunoassay	Solid phase (heterogenous)
immunoassay
Traceability	International Reference Preparation
is not available
Results are traceable to in-house	International Reference Preparation is
not available

6

	Standards
Antigen	Native RNP antigen, purified from
calf thymus	Native RNP antigen, purified from calf
thymus
Sample type	Serum	Serum
Shelf life	One year	One year

Differences
Item	QUANTA Flash RNP	Predicate Device
Detection/
Operating principle	Chemiluminescent immunoassay	Enzyme-linked immunosorbent assay
Solid phase	Paramagnetic microparticles (beads)	96-well plate
Conjugate	Isoluminol conjugated anti-human IgG	HRP conjugated anti-human IgG
Calibration	Lot specific Master Curve + two
calibrators (Sold separately)	RNP ELISA Low Positive
(Included in the kit)
Item	QUANTA Flash Sm Calibrators	Predicate Device
Intended use	For use with the QUANTA Sm
chemiluminescent
immunoassay
(CIA). Each calibrator establishes a
point of reference for the working
curve that is used to determine
Chemiluminescent Unit (CU) values
in the measurement of anti-Sm
antibodies in human serum.	No separate intended use;
calibrators are part of the kit.
Analyte	Anti-Sm antibodies	Anti-Sm antibodies
Method	QUANTA Flash Sm
chemiluminescent immunoassay	QUANTA Lite Sm ELISA
Unit	CU (Chemiluminescent units)
(arbitrary)	Units (arbitrary)
Matrix	Human serum, buffer, stabilizers,
preservative	Human serum, buffer, stabilizers,
preservative
Physico-chemical
characteristics	Liquid, ready to use	Liquid, ready to use
Storage	2-8 °C	2-8 °C
Shelf life	One year	One year
In-use stability	Four calibrations, maximum total 8 hours uncapped onboard the instrument.	Calibrators can be used until the end
of the shelf life when stored
properly

QUANTA Flash Sm Calibrators

QUANTA Flash RNP Calibrators

Item	QUANTA Flash RNP Calibrators	Predicate Device
------	------------------------------	------------------

7

| Intended use | For use with the QUANTA Flash RNP
chemiluminescent
immunoassay
(CIA). Each calibrator establishes a
point of reference for the working
curve that is used to determine
Chemiluminescent Unit (CU) values
in the measurement of anti-RNP
antibodies in serum. | No separate intended use;
calibrators are part of the kit. |
|-------------------------------------|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|------------------------------------------------------------------------------------|
| Analyte | Anti-RNP antibodies | Anti-RNP antibodies |
| Method | QUANTA Flash RNP
chemiluminescent immunoassay | QUANTA Lite RNP ELISA |
| Matrix | Human serum, buffer, stabilizers,
preservative | Human serum, buffer, stabilizers,
preservative |
| Unit | CU (Chemiluminescent units)
(arbitrary) | Units (arbitrary) |
| Physico-chemical
characteristics | Liquid, ready to use | Liquid, ready to use |
| Storage | 2-8 °C | 2-8 °C |
| Shelf life | One year | One year |
| In-use stability | Four calibrations, maximum total 8
hours uncapped onboard the
instrument. | Calibrators can be used until the end
of the shelf life when stored
properly |

QUANTA Flash Sm Controls

Item	QUANTA Flash Sm Controls	Predicate Device
Intended use	Quality control purposes of the
QUANTA Flash Sm
chemiluminescent immunoassay
(CIA) kit.	No separate intended use; controls
are part of the kit.
Analyte	Anti-Sm antibodies	Anti-Sm antibodies
Method	QUANTA Flash Sm
chemiluminescent immunoassay	QUANTA Lite Sm ELISA
Unit	CU (Chemiluminescent units)
(arbitrary)	Units (arbitray)
Matrix	Human serum, buffer, stabilizers,
preservative	Human serum, buffer, stabilizers,
preservative
Physico-chemical
characteristics	Liquid, ready to use	Liquid, ready to use
Levels	2 (negative and positive)	2 (ELISA negative, high positive)
Storage	2-8 °C	2-8 °C
Shelf life	One year	One year
In-use stability	15 uses, with a maximum time of 10
minutes onboard the instrument per
use, or 2 ½ hours, total.	Controls can be used until the end
of the shelf life when stored
properly

:

.

8

Item	QUANTA Flash RNP Controls	Predicate Device
Intended use	Quality control purposes of the
QUANTA Flash RNP
chemiluminescent immunoassay
(CIA) kit.	No separate intended use; controls
are part of the kit.
Analyte	Anti-RNP antibodies	Anti-RNP antibodies
Method	QUANTA Flash RNP
chemiluminescent immunoassay	QUANTA Lite RNP ELISA
Matrix	Human serum, buffer, stabilizers,
preservative	Human serum, buffer, stabilizers,
preservative
Unit	CU (Chemiluminescent units)
(arbitrary)	Units (arbitray)
Physico-chemical
characteristics	Liquid, ready to use	Liquid, ready to use
Levels	2 (negative and positive)	2 (ELISA negative, high positive)
Storage	2-8 °C	2-8 °C
Shelf life	One year	One year
In-use stability	15 uses, with a maximum time of 10
minutes onboard the instrument per
use, or 2 ½ hours, total.	Controls can be used until the end
of the shelf life when stored
properly

OUANTA Flash RNP Controls

Value assignment and traceability of Calibrators and Controls

The QUANTA Flash Sm and RNP Calibrators and Controls are manufactured by diluting human serum that contains high titer of anti-RNP antibodies with a buffer containing stabilizers and preservative. The human serum is obtained from commercial sources and it is tested for markers of infectious substances.

The target CU is achieved through trial dilutions on small scale. Once a dilution is selected, the Calibrators and Control are bulked, tested, and adjusted. Upon completion of the manufacturing process, the Calibrators and Controls are tested on at least two instruments, on at least two lots of reagent cartridge, in replicates of 10 to determine final value assignment.

There are currently no recognized international standards for the measurement of anti-RNP antibodies.

Calibrator and Control values are directly traceable to in-house Standards that are used to create the Master Curve for the QUANTA Flash Sm and QUANTA Flash RNP assays.

Performance characteristics

Precision

The precision of the QUANTA Flash Sm assay was evaluated on 5 samples containing various concentrations of Sm antibodies in accordance with CLSI EP5-A2, Evaluation of Precision Performance of Quantitative Measurement Procedures - Approved Guideline: samples were run in duplicates, twice a day, for at least 20 days. Data were analyzed with the Analyse-it for Excel method evaluation software,

9

| | | | | Within-Run
Precision
(repeatability) | | Between-Run | | Between-Day
Precision | | Total Precision | |
|--------------|----|--------------|-----|--------------------------------------------|-----|-------------|-----|--------------------------|-----|-----------------|--|
| Sample
ID | N | Mean
(CU) | SD | %CV | SD | %CV | SD | %CV | SD | %CV | |
| 1 | 80 | 13.1 | 0.9 | 6.5 | 0.2 | 1.5 | 0.7 | 5.6 | 1.1 | 8.7 | |
| 2 | 80 | 22.0 | 2.1 | 9.7 | 1.0 | 4.6 | 0.9 | 4.2 | 2.5 | 11.5 | |
| 3 | 84 | 93.0 | 6.0 | 6.4 | 0.5 | 0.5 | 4.8 | 5.2 | 7.7 | 8.3 | |

16.8

ਰੇ ਰੇ

7.1

2.9

6.4

19.7

2.7

5.8

27.3

28.5

11.5

8.4

and within run, between run, between day and total precisions are summarized in the Table below. All %CV values were within the acceptance limit, 15%.

The precision of the QUANTA Flash RNP assay was evaluated on 7 samples containing various concentrations of RNP antibodies in accordance with CLSI EP5-A2, Evaluation of Precision Performance of Quantitative Measurement Procedures - Approved Guideline: samples were run in duplicates, twice a day, for at least 20 days. Data were analyzed with the Analyse-it for Excel method evaluation software, and within run, between run, between day and total precision are summarized in the Table below. All %CV values were within the acceptance limit, 15%.

| | | | | Within-Run
Precision
(repeatability) | | Between-Run
Precision | | Between-Day
Precision | | Total Precision | |
|--------------|----|-----------|--|--------------------------------------------|-----|--------------------------|-----|--------------------------|-----|-----------------|------|
| Sample
ID | N | Mean (CU) | | SD | %CV | SD | %CV | SD | %CV | SD | %CV |
| 1 | 92 | 6.7 | | 0.2 | 3.5 | 0.0 | 0.0 | 0.4 | 5.3 | 0.4 | 6.4 |
| 2 | 84 | 24.8 | | 0.9 | 3.7 | 0.0 | 0.0 | 0.8 | 3.0 | 1.2 | 4.8 |
| 3 | 88 | 31.6 | | 1.1 | 3.4 | 0.7 | 2.2 | 2.7 | 8.5 | 3.0 | 9.4 |
| 4 | 92 | 120.7 | | 4.9 | 4.1 | 4.5 | 3.7 | 4.5 | 3.7 | 8.0 | 6.6 |
| 5 | 88 | 218.6 | | 9.3 | 4.2 | 7.8 | 3.6 | 20.3 | 9.3 | 23.6 | 10.8 |
| 6 | 88 | 319.5 | | 12.0 | 3.7 | 14.0 | 4.4 | 11.6 | 3.6 | 21.8 | 6.8 |
| 7 | 96 | 409.6 | | 19.6 | 4.8 | 18.2 | 4.5 | 23.3 | 5.7 | 35.5 | 8.7 |

Reproducibility

4

5

88

84

237.7

338.6

20.6

18.1

8.7

5.3

Three samples for Sm and RNP each were tested on two different regent lots, using two different lots of Calibrators, by two operators. Samples were run in quadruplicates, two times a day, for 10 days, to generate 80 data points. Data were analyzed with the Analyse-it for Excel method evaluation software, and within run, between reagent lots, between calibrator lots, between operators and total precision were calculated and the results are summarized in the Tables below. All %CV values were within the acceptance limit, 15%.

10

QUANTA Flash Sm

| | | | Within Run | | Between | Reagent Lots | | Between
Calibrator
Lots | | Between
Operators | | Total |
|--------------|--------------|----------------------------|------------|-----------|------------|--------------|------------|-------------------------------|------------|----------------------|------------|-----------|
| Sample | Meán
(CU) | Number
of
replicates | SD
(CU) | CV
(%) | SD
(CU) | CV
(%) | SD
(CU) | CV
(%) | SD
(CU) | CV
(%) | SD
(CU) | CV
(%) |
| Sample
#1 | 15.2 | 80 | 0.4 | 2.8 | 1.4 | 9.4 | 1.1 | 6.9 | 1.1 | 6.9 | 1.1 | 7.0 |
| Sample
#2 | 23.3 | 80 | 0.6 | 2.4 | 2.5 | 10.7 | 1.8 | 7.6 | 1.7 | 7.3 | 1.8 | 7.7 |
| Sample
#3 | 157.3 | 80 | 5.5 | 3.5 | 15.8 | 10.1 | 15.1 | 9.6 | 13.1 | 8.4 | 13.0 | 8.3 |

QUANTA Flash RNP

| | | | Within Run | | Between
Reagent Lots | | Between
Calibrator
Lots | | Between
Operators | | Total | | |
|--------------|--------------|----------------------------|------------|-----------|-------------------------|-----------|-------------------------------|-----------|----------------------|-----------|------------|-----------|--|
| Sample | Mean
(CU) | Number
of
replicates | SD
(CU) | CV
(%) | SD
(CU) | CV
(%) | SD
(CU) | CV
(%) | SD
(CU) | CV
(%) | SD
(CU) | CV
(%) | |
| Sample
#1 | 15.3 | 80 | 0.4 | 2.9 | 0.5 | 3.1 | 0.5 | 3.3 | 0.3 | 1.8 | 0.4 | 2.8 | |
| Sample
#2 | 23.1 | 80 | 0.5 | 2.3 | 0.7 | 2.8 | 0.7 | 3.1 | 0.4 | 1.7 | 0.6 | 2.6 | |
| Sample
#3 | 173.1 | 80 | 5.4 | 3.1 | 6.2 | 3.6 | 5.1 | 2.9 | 3.1 | 1.8 | 5.1 | 2.9 | |

Limit of Blank, Limit of Detection

QUANTA Flash Sm:

The Limit of Detection (LoD) of the QUANTA Flash Sm assay is 803 RLU, which is below the analytical measuring range of the assay. It was determined consistent with CLSI EP17-A guideline with proportions of false positives (alpha) less than 5% and false negatives (beta) less than 5%; based on 140 determinations, with 60 measurements on blank samples and 80 measurements of low level samples. The LoB is 540 RLU. Because the curve ends at 3.3 RLU (which is equal to approximately 2430 RLU), it was not possible to calculate the LoB (540 RLU) and LoD ( 803 RLU) in CUs, only in RLUs (instrument signal).

QUANTA Flash RNP:

The Limit of Detection (LoD) of the QUANTA Flash RNP assay is 1128.8 RLU, which is below the analytical measuring range of the assay. It was determined consistent with CLSI EP17-A guideline with proportions

11

of false positives (alpha) less than 5% and false negatives (beta) less than 5%; based on 140 determinations, with 60 measurements on blank samples and 80 measurements of low level samples. The LoB is 713 RLU. Because the curve ends at 3.5 RLU (which is equal to approximately 3450 RLU), it was not possible to calculate the LoB (713 RLU) and LoD (1128.8 RLU) in CUs, only in RLUs (instrument signal).

Analytical Measuring Range

QUANTA Flash Sm:

The analytical measuring range (AMR) of the assay (determined by the lowest and highest points of the Master Curve) is 3.3 CU to 693.5 CU, which corresponds to the linear range of the assay. If a patient result is less than 3.3 CU, the BIO-FLASH system will report it as "693.5 CU". This is considered a positive result. The BIO-FLASH software has an Auto-Rerun option available (see description and validation in the next paragraph). If this option is selected, the instrument will automatically rerun any sample that has a result of >693.5 CU after additional 10 fold dilution, thereby bringing the measured value within the AMR.

The linearity of the AMR was evaluated by a study according to CLSI EP6-A, Evaluation of the Linearity of Quantitative Measurement Procedures: A Statistical Approach; Approved Guideline. Five serum samples with various Sm antibody concentrations were diluted with a low negative serum to obtain values that covered a range approximately 20% wider than the AMR. All specimens showed dilution linearity individually, and the combined data yielded the following results with linear regression:

Sample	Slope (95% Cl)	Y-intercept (95% Cl)	R2
All Samples (n=5)	0.99 (0.97 to 1.00)	0.81 (-3.2 to 4.8)	0.99

QUANTA Flash RNP:

The analytical measuring range (AMR) of the assay (determined by the lowest and highest points of the Master Curve) is 3.5 CU to 643.8 CU, which corresponds to the linear range of the assay. If a patient result is less than 3.5 CU, the BIO-FLASH system will report it as "643.8 CU". This is considered a positive result. The BIO-FLASH software has an Auto-Rerun option available (see description and validation in the next paragraph). If this option is selected, the instrument will automatically rerun any sample that has a result of >643.8 CU after additional 10 fold dilution, thereby bringing the measured value within the AMR.

The linearity of the AMR was evaluated by a study according to CLSI EP6-A, Evaluation of the Linearity of Quantitative Measurement Procedures: A Statistical Approach; Approved Guideline. Five serum samples with various anti-RNP concentrations were diluted with a low negative serum to obtain values that covered a range approximately 20% wider than the AMR. All six specimens showed dilution linearity individually, and the combined data yielded the following results with linear regression:

Sample	Slope (95% CI)	Y-intercept (95% CI)	R2
All Samples (n=5)	1.00 (1.00 to 1.02)	-5.17 (-8.27 to -2.07)	1.00

12

Auto-rerun function

The BIO-FLASH software has an Auto-rerun option available. If this option is selected, the instrument will automatically rerun any sample that has a result of >693.5 CU for Sm and > 643.8 CU for RNP after additional 10 fold dilution, thereby bringing the measured value within the AMR. The final result will be calculated by the software. As the highest value that can be measured is 693.5 CU for Sm and 643.8 CU for RNP, the highest value that can be reported is 6935 CU and 6438 CU, respectively.

To validate the Auto-rerun function, five (Sm) and six (RNP) high positive specimens with results above the analytical measuring range were selected for the Sm and the RNP assays, respectively. The samples were run with the Auto-rerun function enabled on the BIO-FLASH. Then the specimens were manually diluted the same way as it happens in the Auto-rerun function (10 fold dilution), and tested on the BIO-FLASH. The results were within the analytical measuring range after auto-rerun or manual dilution for all specimens. The % recovery values for results obtained with the auto-rerun results compared to the results obtained by manual dilution were between 87.6% and 97.7% for Sm (average 91.5%), and between 82.5% and 102.2% for RNP (average 96.3%), respectively (within the ± 20% acceptance limit).

Interference

QUANTA Flash Sm:

The interference study was performed according to CLSI EP07-A2, Interference Testing in Clinical Chemistry; Approved Guideline - Second Edition. Three specimens were tested (negative: 16.3 CU); around the cutoff weak positive: 28.6 CU; high positive: 190.7 CU). Interfering substances were spiked into every specimen at three different concentrations in 10% of total specimen volume, and the resulting samples were assessed in triplicates with the Sm assay. Recovery of the unit values was calculated compared to control samples spiked with the same volume of diluents. Acceptance criteria for the interference studies were 85% - 115% recovery, or ± 4 CU difference, whichever is greater.

No interference was detected with bilirubin up to 10 mg/dL (recovery: 103% to 108% or within ± 4 CU), hemoglobin up to 200 mg/dL (recovery: 97% to 108% or within ± 4 CU), triglycerides up to 1000 mg/dL (recovery: 105% to 111% or within ± 4 CU), cholesterol up to 224.3 mg/dL (recovery: 105% to 111% or within ± 4 CU), and RF IgM up to 500 IU/mL (recovery: 85% to 95% or within ± 4 CU).

QUANTA Flash RNP:

The interference study was performed according to CLSI EP07-A2, Interference Testing in Clinical Chemistry; Approved Guideline - Second Edition. Three specimens were tested (negative: 8.8 CU; weak positive: 55.5 CU; high positive: 280.3). Interfering substances were spiked into every specimen at three different concentrations in 10% of total specimen volume, and the resulting samples were assessed in triplicates with the RNP assay. Recovery of the unit values was calculated compared to control samples spiked with the same volume of diluents. Acceptance criteria for the interference studies were 85% -115% recovery, or ± 4 CU difference, whichever is greater.

No interference was detected with bilirubin up to 10 mg/dL (recovery: 96% to 112%), hemoglobin up to 200 mg/dL (recovery: 88% to 108%), triglycerides up to 1000 mg/dL (recovery: 86% to 109%), cholesterol up to 224.3 mg/dL (recovery: 86% to 109%), and RF IgM up to 500 IU/mL (recovery: 88% to 113%).

13

High concentration hook effect

To assess hook effect, the measurement signal (relative light units, RLU) was examined for the samples that were used to validate the Auto-rerun function (five Sm and six RNP high positive specimens), before and after automatic or manual dilution. All sera produced significantly higher RLU values (above the AMR) when used "as is" compared to the manually or automatically diluted ones, thereby confirming that high positive specimens above the analytical measuring range do not show hook effect up to 2429 CU in the Sm assay and up to 3140 CU in the RNP assay (the highest concentrations tested).

Cross-reactivity

QUANTA Flash Sm:

To test potential cross-reactivity with autoantibodies and infection-induced antibodies, 233 patient samples with various antibodies to autoimmune or infectious disease markers were tested. Three samples from patients with scleroderma, one sample with HIV antibodies, and one sample with MCTD/primary biliary cirrhosis (PBC) overlap were positive on the QUANTA Flash Sm. All other serum samples were negative:

Patient Group	N	Positive	Percent Positive
Autoimmune liver disease	2	0	0%
Viral hepatitis	19	0	0%
Scleroderma	74	3	4.1%
Sjögren's Syndrome	5	0	0%
Rheumatoid arthritis	70	0	0%
Systemic rheumatic disease, other	53	1	1.9%
Infectious disease (HIV + syphilis)	10	1	10%
Total	233	5	2.1%

QUANTA Flash RNP:

To test potential cross-reactivity with autoantibodies and infection-induced antibodies, 246 patient samples were tested from patients with infectious diseases and connective tissue diseases. Eight samples from patients with scleroderma, two samples from patients with RA, one sample from a patient with PM/DM, and one sample with HIV antibodies were positive on the QUANTA Flash RNP. As anti-RNP antibodies can sometimes be detected in patients with systemic sclerosis, and as all of these scleroderma samples were positive on the predicate RNP ELISA, too, it is highly possible that those samples are true positives, and not cross-reacting samples. All other serum samples were negative:

Patient Group	N	Positive	Percent Positive
Autoimmune liver disease	2	0	0%
Viral hepatitis	20	0	0%
Scleroderma	76	8	10.5%
Sjögren's Syndrome	6	0	0%
Rheumatoid arthritis	70	2	2.9%

14

Systemic rheumatic disease, other	48	0	0%
Poly- and Dermatomyositis	14	1	7.1
Infectious disease (HIV + syphilis)	10	1	10%
Total	246	12	4.9%

Stability

Shelf life

To establish the initial claim for shelf life, accelerated stability studies were performed.

Accelerated stability testing was performed on each of the following sealed components in the QUANTA Flash Sm and RNP to establish initial stability claim: the beads, the two calibrators, and the negative and positive controls. Each week a new sealed component was placed in the incubator, and all components were tested at the end of the experiment together with the one that was stored at 5º ± 3ºC. The recovery of the measured values was calculated for each time point (compared to those obtained with 5º ± 3ºC stored reagent). All calculations were performed by comparing results of sealed components stored at 5º ± 3ºC (control) to those stored at 37º ± 3ºC (test) for 1, 2, 3, and 4 weeks, where one week is equal to six months at 5º ± 3ºC. Linear regression analysis was performed between recovery values and the number of days in those cases when at least 3 data points were available at each time point; otherwise, individual data points were analyzed.

Acceptance criteria for one year preliminary expiration dating were:

- Controls and Calibrators:

a) if regression analysis is used, the lower 95% Cl interval of the regression line is > 90% at 2 weeks, and no individual data point has ≤ 80% recovery,

b) If individual data points are analyzed, recovery values are > 90% at day 14.

Microparticles:

with regression analysis, the lower 95% Cl interval of the regression line is ≥ 85% at 2 weeks, and no individual data point has ≤ 75% recovery.

Sm beads, RNP beads, Sm Calibrators, RNP Calibrators, Sm Controls and RNP Controls each fulfilled the acceptance criteria above, so one year expiration dating was assigned to each component.

In-use (onboard) stability

Sm Calibrators

During assessing on-board stability, Calibrators were placed, uncapped, onboard the instrument, and calibration was performed altogether five times, then a panel of characterized patient specimens were run on each calibration curve.

Acceptance criteria were: Calibrators are considered stable if all five calibrations performed in the 8.5 hour period are successful, and Calibrator RLU recovery values are between 90% and 110% compared to the first use.

15

A total of 5 successful calibrations were performed over an 8.5 hour period. Calibrator RLU values remained within the 90-110% range. Moreover, all patient panel samples ran within their expected range. This supports the claim that calibrators can be used for up to 4 calibrations over an 8 hour period.

Sm Controls

During assessing on-board stability, Controls were assayed twice each day over 10 days, for a total of 20 runs. The controls were left uncapped, onboard the instrument for 15 minutes per run. When not in use, the controls were capped, and stored at 2-8°C.

Acceptance criteria: Controls are considered stable when all replicates run within their established range, moreover, and the linear regression line obtained by plotting %recovery values against the number of runs stays between 85% and 115% at day run 15.

Both controls ran within their respective acceptable range for all 20 runs, resulting in a %CV of 5.9% for the Negative Control and 4.8% for the Positive Control. These results support the claim that controls can be used for 15 times, up to 2.5 hours total.

Sm Reagent Cartridge

To determine the in-use stability of the QUANTA Flash Sm reagent pack, three lots of cartridges were tested by using five serum specimens (with different reactivity levels). The specimens were tested periodically up to 48 days, twice a day. Recoveries were calculated compared to the day zero average values, and linear regression analysis was performed. The claim was established using the following criteria (using the one that is fulfilled first):

a)The stability claim is established at the day where the lower 95% confidence interval of the regression line reaches 85% recovery, or

b) When 2% or more of the recovery data is ≤ 75%.

The onboard stability results of the three lots are the following: RP0005: 33 days RP0006: 40 days 121005: 39 days

Using these criteria, the in-use (onboard) stability of the Sm reagent cartridge was set at 33 days.

RNP Calibrators

During assessing on-board stability, Calibrators were placed, uncapped, onboard the instrument, and calibration was performed altogether five times, then a panel of characterized patient specimens were run on each calibration curve.

Acceptance criteria were: Calibrators are considered stable if all five calibrations performed in the 8.5 hour period are successful, and Calibrator RLU recovery values are between 90% and 110% compared to the first use.

16

A total of 5 successful calibrations were performed over an 8.5 hour period. Calibrator RLU values remained within the 90-110% range. Moreover, all patient panel samples ran within their expected range. This supports the claim that calibrators can be used for up to 4 calibrations over an 8 hour period.

RNP Controls

During assessing on-board stability, Controls were assayed twice each day over 10 days, for a total of 20 runs. The controls were left uncapped, onboard the instrument for 15 minutes per run. When not in use; the controls were capped, and stored at 2-8℃.

Acceptance criteria: Controls are considered stable when all replicates run within their established range, moreover, and the linear regression line obtained by plotting %recovery values against the number of runs stays between 85% and 115% at day run 15.

Both controls ran within their respective acceptable range for all 20 runs, resulting in a %CV of 5.9% for the Negative Control and 6.9% for the Positive Control. These results support the claim that controls can be used for 15 times, up to 2.5 hours total.

RNP Reagent Cartridge

To determine the in-use stability of the QUANTA Flash RNP reagent pack, three lots of cartridges were tested by using four to eight serum specimens (with different reactivity levels). The specimens were tested periodically up to 41 days, twice a day. Recoveries were calculated compared to the day zero average values, and linear regression analysis was performed. The claim was established using the following criteria (using the one that is fulfilled first):

alThe stability claim is established at the day where the lower 95% confidence interval of the regression line reaches 85% recovery, or

b) When 2% or more of the recovery data is ≤ 75%.

The onboard stability results of the three lots are the following: 111001: 28 days 121002: 28 days 121003: 31 days

Using these criteria, the in-use (onboard) stability of the RNP reagent cartridge was set at 28 days.

The Stability Claims are tabulated below for easy review. The 1 year shelf life begins from the date of manufacture.

Sm stability claims

Product	Unopened shelf life	After being opened
QUANTA Flash Sm
Reagent Cartridge	Until the expiration
date on the label
(currently 1 year).	33 days (onboard only – cannot be
resealed).

17

| Sm Calibrator 1 and 2 | Until the expiration
date on the label
(currently 1 year). | 4 uses, up to 8 hours onboard total |
|---------------------------------------------|------------------------------------------------------------------|----------------------------------------|
| Sm Negative Control
and Positive Control | Until the expiration
date on the label
(currently 1 year). | 15 uses, up to 2.5 hours onboard total |

RNP stability claims

Product	Unopened shelf life	After being opened
QUANTA Flash RNP
Reagent Cartridge	Until the expiration
date on the label
(currently 1 year).	28 days (onboard only – cannot be
resealed).
RNP Calibrator 1 and 2	Until the expiration
date on the label
(currently 1 year).	4 uses, up to 8 hours onboard total
RNP Negative Control
and Positive Control	Until the expiration
date on the label
(currently 1 year).	15 uses, up to 2.5 hours onboard total

Cut-off, reference range

| QUANTA Flash Sm: | Negative
Positive |