The BioPlex®2200 Antiphospholipid Syndrome (APLS) IgM kit is a multiplex flow immunoassav intended for the semi-quantitative detection of IgM antibodies to Cardiolipin (CL) and Beta-2 Glycoprotein I (B2GPI) in human serum and plasma (lithium heparin, sodium heparin, and sodium citrate). In conjunction with clinical findings, the test system is used as an aid in the diagnosis of primary Antiphospholipid Syndrome (APS) and those secondary to systemic lupus erythematosus (SLE) or SLE-like disorders.

The BioPlex 2200 APLS IgM kit is intended for use with the Bio-Rad BioPlex 2200 System.

The BioPlex 2200 Antiphospholipid Syndrome (APLS) IgM Calibrator Set is intended for the calibration of the corresponding BioPlex 2200 APLS IgM Reagent Pack.

The BioPlex 2200 Antiphospholipid Syndrome (APLS) IgM Control Set is intended for use as an assayed quality control to monitor the overall performance of the BioPlex 2200 Instrument and the corresponding BioPlex 2200 APLS IgM Reagent Pack in the clinical laboratory. The performance of the BioPlex 2200 APLS IgM Control Set has not been established with any other Antiphospholipid assay.

Device Description

BioPlex® 2200 APLS IgM kit includes the following components:

o One (1) 10 mL vial of Bead Set containing two different populations of dyed beads coated with Cardiolipin (CL) and Beta-2-Glycoprotein I (B2GPI). an Internal Standard bead (ISB). a Serum Verification bead (SVB), and a Reagent Blank bead (RBB) in a MOPS (3-[N-Morpholino] propanesulfonic acid) buffer supplemented with glycerol and protein stabilizers (porcine), and ProClin 300 (< 0.3%), sodium benzoate (≤ 0.1%) and sodium azide (< 0.1%) as preservatives.
One (1) 5 mL vial of Conjugate containing phycoervthrin conjugated murine . monoclonal anti-human IgM and phycoerythrin conjugated murine monoclonal anti-human FXIII in MOPS (3-[N-Morpholino] propanesulfonic acid) buffer supplemented with protein stabilizers (bovine), and ProClin 300 (< 0.3%), sodium benzoate (≤ 0.1%) and sodium azide (< 0.1%) as preservatives.
One (1) 10 mL vial of Sample Diluent containing buffer with protein stabilizers . (bovine and murine), and ProClin 300 (< 0.3%). sodium benzoate (≤ 0.1%) and sodium azide (< 0.1%) as preservatives.

BioPlex® 2200 APLS IgM Calibrator set contains seven 0.5 mL vials of human antibodies to CL or B2GPI in a human serum matrix made from defibrinated plasma with ProClin 300 (≤ 0.3%), sodium benzoate (< 0.1%) and sodium azide (< 0.1%) as preservatives.

BioPlex® 2200 APLS [gM Control set contains four 1.5-mL vials of Positive controls of human antibodies to CL or ß2GPI and two vials of Negative Controls in a human serum matrix made from defibrinated plasma; and, in a human serum matrix made from defibrinated plasma with ProClin 300 (≤0.3%), sodium benzoate (≤ 0.1%) and sodium azide (< 0.1%) as preservatives.

Additional materials required but not supplied include BioPlex® 2200 Sheath Fluid containing Phosphate Buffered Saline (PBS) with ProClin® 300 (0.03%) and sodium azide (<0.1%) as preservatives; and BioPlex® 2200 Wash Solution containing Phosphate Buffered Saline (PBS) and Tween 20 with ProClin® 300 (<0.03%) and sodium azide (<0.1%) as preservatives.

AI/ML Overview

Here's a summary of the acceptance criteria and study information for the BioPlex® 2200 Antiphospholipid Syndrome (APLS) IgM kit, based on the provided text:

Acceptance Criteria and Device Performance

Acceptance Criteria Category	Specific Criteria	Reported Device Performance
Precision/Reproducibility	Total precision for Anti-Cardiolipin IgM (serum & heparin)	%CV ≤ 8.1% (High Positive serum)
	Total precision for Anti-β2GPI IgM (serum & heparin)	%CV ≤ 8.0% (High Positive serum)
Lot-to-Lot Reproducibility	Not explicitly stated but implied by detailed results.	For Anti-Cardiolipin IgM, Total %CV ranged from 5.6% to 11.8%. For Anti-β2GPI IgM, Total %CV ranged from 6.8% to 14.1%.
Linearity/Assay Reportable Range	Regression parameters (slope, intercept, r²) for linearity study. Reportable range.	Slopes near 1.00, intercepts near 0.00, r² > 0.99. Reportable range: Anti-Cardiolipin IgM (0.2 to 112 MPL-U/mL), Anti-β2GPI IgM (0.2 to 112 U/mL).
Detection Limit (LoQ, LoD, LoB)	Specific values for LoQ, LoD, LoB.	Anti-Cardiolipin IgM: LoQ: 0.2, LoD: 0.13, LoB: 0.0974 MPL-U/mL. Anti-β2GPI IgM: LoQ: 0.2, LoD: 0.13, LoB: 0.1044 U/mL.
Analytical Specificity (Interference)	No interference observed with specified substances at maximum levels.	No interference observed with: Hemoglobin (≤500 mg/dL), Bilirubin (unconjugated ≤20 mg/dL, conjugated ≤30 mg/dL), Triglycerides (≤3300 mg/dL), Total Protein (≤12 g/dL), Cholesterol (≤500 mg/dL), Red Blood Cells (≤0.4% (v/v)), Gamma-Globulin (≤6 g/dL), Beta-Carotene (≤0.6 mg/dL), Ascorbic Acid (≤3 mg/dL), Heparin Lithium (≤8000 units/dL), Heparin Sodium (≤8000 units/dL), Sodium Citrate (<1000 mg/dL).
Cross-Reactivity	Expected positivity rates for various disease states.	Positivity rates varied by disease state and IgM type (e.g., Systemic Lupus Erythematosus: 5.9% for both, Scleroderma: 15.0% for both, Syphilis: 0% for both).
Assay Cut-off	Cutoff value established to achieve 99% clinical specificity in a normal healthy population.	Cutoff of 20.0 MPL-U/mL for anti-CL IgM and 20 U/mL for anti-B2GPI IgM.
Method Comparison (Positive Agreement)	Not explicitly stated but calculated as performance against predicate.	Anti-Cardiolipin IgM: Positive Agreement = 73.3% (77/105) (95% CI: 64.2 - 80.9%).
Method Comparison (Negative Agreement)	Not explicitly stated but calculated as performance against predicate.	Anti-Cardiolipin IgM: Negative Agreement = 99.8% (403/404) (95% CI: 98.6 - 100%).
Method Comparison (Total Agreement)	Not explicitly stated but calculated as performance against predicate.	Anti-Cardiolipin IgM: Total Agreement = 94.3% (480/509) (95% CI: 91.9 - 96.0%).
Method Comparison (Positive Agreement)	Not explicitly stated but calculated as performance against predicate.	Anti-β2GPI IgM: Positive Agreement = 95.2% (79/83) (95% CI: 88.3 - 98.1%).
Method Comparison (Negative Agreement)	Not explicitly stated but calculated as performance against predicate.	Anti-β2GPI IgM: Negative Agreement = 94.0% (233/248) (95% CI: 90.3 - 96.3%).
Method Comparison (Total Agreement)	Not explicitly stated but calculated as performance against predicate.	Anti-β2GPI IgM: Total Agreement = 94.3% (312/331) (95% CI: 91.2 - 96.3%).
Matrix Comparison	Slope of 1.00 ± 0.2, y-intercept of 0.0 ± 6.0, r between 0.980 and 1.000 for regression of serum vs. plasma.	All tested matrix comparisons (Lithium Heparin vs. Serum, Sodium Heparin vs. Serum, Sodium Citrate vs. Serum) fell within the specified ranges for slope, intercept, and r for both aCL IgM and aβ2GPI IgMassays.
Clinical Sensitivity	Not explicitly stated but implied by reported values.	Anti-Cardiolipin IgM: 33.7% (67/199) (95% CI: 27.5 - 40.5%). Anti-β2GPI IgM: 40.2% (80/199) (95% CI: 33.6-47.1%).
Clinical Specificity	Not explicitly stated but implied by reported values.	Anti-Cardiolipin IgM: 96.8% (335/346) (95% CI: 94.4 - 98.2%). Anti-β2GPI IgM: 96.0% (332/346) (95% CI: 93.3 - 97.6%).

Study Details

Sample size used for the test set and the data provenance:
- Precision/Reproducibility (CLSI EP5-A2):
  - Serum and heparinized plasma panels: Assayed in replicate twice daily over 20 days (n=80) for most samples, and over 10 days (n=40) for mid-negative samples.
  - Data provenance: Not explicitly stated (e.g., country of origin). The study seems to be prospective in nature, conducted specifically for device validation.
- Precision/Reproducibility (CLSI EP15-A2):
  - Serum panel: Assayed in 4 replicates per run, one run per day over 5 days (n=20).
  - Data provenance: Not explicitly stated. Prospective.
- Lot-to-Lot Reproducibility:
  - Serum samples: Replicates of 10 for two runs. Total of 60 points for each patient serum sample.
  - Data provenance: Not explicitly stated. Prospective.
- Linearity/Assay Reportable Range:
  - Six positive patient samples.
  - Data provenance: Not explicitly stated. Prospective.
- Analytical Specificity (Interference):
  - Not specified how many samples were tested for each substance.
  - Data provenance: Not explicitly stated. Prospective.
- Cross-Reactivity:
  - Number of samples varied by disease state (e.g., Systemic Lupus Erythematosus: 34, Scleroderma: 20, Syphilis: 15).
  - Data provenance: Samples from individuals with known disease states. Implies retrospective collection of known disease state samples.
- Assay Cut-off Determination:
  - 103 samples from patients diagnosed as primary or secondary APS.
  - 123 samples from non-APS or other rheumatic disease control donors.
  - 208 samples from apparently healthy donors.
  - Data provenance: Implies retrospective collection for diagnosed patients and prospective for healthy donors.
- Method Comparison with Predicate Device:
  - 199 patients diagnosed with primary or secondary APS.
  - 346 patients with other rheumatic or non-APS disease.
  - Data provenance: Implies retrospective collection of diagnosed patient samples and controls.
- Matrix Comparison:
  - 38 matched sets of serum, heparin, and citrate plasma samples.
  - Data provenance: Samples drawn from the same donor. Prospective for matched samples.
- Clinical Sensitivity and Specificity:
  - 199 diagnosed primary or secondary APS patients.
  - 346 non-APS disease control patients.
  - Data provenance: Implies retrospective collection of diagnosed patient samples and controls.
- Expected Values/Reference Range:
  - 300 samples from apparently healthy donors (132 males, 168 females).
  - Data provenance: Apparently healthy donors are usually prospectively collected.
Number of experts used to establish the ground truth for the test set and the qualifications of those experts:
- The document does not explicitly state the number of experts or their qualifications used to establish ground truth for the test set.
- For the clinical sensitivity and specificity studies, patients were "diagnosed primary or secondary APS patients" or "non-APS disease control patients." It's implied that these diagnoses were established through standard clinical practice, likely involving medical specialists, but specific details on the number or qualifications of clinicians involved in determining these diagnoses are not provided.
- The assay cutoff determination references "clinical diagnosis as the standard" and the "International Consensus Statement on an Update of the Classification Criteria for Definite Antiphospholipid Syndrome (APS)" for establishing the 99th percentile of a normal healthy population. While this suggests reliance on expert consensus criteria, it doesn't specify experts involved in this specific study's ground truth establishment.
Adjudication method (e.g., 2+1, 3+1, none) for the test set:
- The document does not mention an adjudication method for the test set, as this is a diagnostic assay measuring biomarkers, not an imaging device requiring human reader interpretation and consensus for ground truth. The "ground truth" for clinical performance is based on established clinical diagnoses.
If a multi-reader multi-case (MRMC) comparative effectiveness study was done, If so, what was the effect size of how much human readers improve with AI vs without AI assistance:
- No, a multi-reader multi-case (MRMC) comparative effectiveness study was not conducted. This is an in vitro diagnostic (IVD) immunoassay that directly detects antibodies, not an AI-assisted diagnostic tool for human readers. Therefore, the concept of "human readers improve with AI vs without AI assistance" is not applicable.
If a standalone (i.e. algorithm only without human-in-the loop performance) was done:
- Yes, the performance presented for the BioPlex® 2200 APLS IgM kit is its standalone performance. This device is a semi-quantitative immunoassay that produces a numerical result and a positive/negative interpretation based on a pre-defined cutoff. Its performance characteristics (precision, linearity, detection limits, clinical sensitivity/specificity) are evaluated for the device itself, without a "human-in-the-loop" component for result interpretation.
The type of ground truth used (expert consensus, pathology, outcomes data, etc):
- The ground truth for most clinical performance evaluations (e.g., clinical sensitivity and specificity, assay cut-off) was based on clinical diagnosis (patients diagnosed with primary or secondary APS vs. non-APS disease controls / apparently healthy donors). This clinical diagnosis would inherently involve a combination of clinical findings, potentially other laboratory tests, and ultimately, a physician's or expert's determination.
- For analytical performance characteristics (precision, linearity, detection limits, interference), ground truth is established through controlled laboratory experiments and reference methods, not clinical diagnosis in the same way.
The sample size for the training set:
- The document does not explicitly describe a separate "training set" in the context of machine learning. For an IVD immunoassay, the concept of "training" typically refers to the process of developing and optimizing the assay parameters, including reagent concentrations, reaction conditions, and calibrator assignments. This process would use various samples, but they are not typically referred to as a "training set" in the same way as in AI/ML development. The text describes calibrator assignment, which is part of this assay optimization process.
How the ground truth for the training set was established:
- As mentioned above, there isn't an explicit "training set" in the AI/ML sense. For the development and calibration of the assay:
  - Calibrator Assignment: "Calibrator assignment is established for matched lots of BioPlex® 2200 APLS IgM kit and calibrators using a master set of calibrators as reference and replicate analyses on multiple BioPlex® 2200 instruments." This implies a meticulous, data-driven process using a master reference.
  - Assay Cut-off: The cut-off was determined using "concordance testing and Receiver Operator Characteristic (ROC) analysis using the clinical diagnosis as the standard" on a clinical cohort (103 APS, 123 non-APS controls) and later confirmed with 208 apparently healthy donors. This establishment of the cutoff value aligns with how a "ground truth" might be used in an IVD context to define the diagnostic threshold.

Summary

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BioPlex® 2200 Antiphospholipid Syndrome (APLS) IgM 510(k) Summary

Bio-Rad Laboratories hereby submits this 510(k) in accordance with the requirements of SMDA 1990 and 21 CFR 807.92. This summary of 510(k) safety and effectiveness information provides detail as a basis for a determination of substantial equivalence for the BioPlex® 2200 APLS IgM kit.

510(k) Number: K130528

Summary Preparation Date: October 18, 2013

OCT 2 1 2013

Applicant: Bio-Rad Laboratories

Contact:

Juang Wang Regulatory Affairs Representative 5500 East Second Street Benicia CA 94510 Ph: (510) 741-4609 Fax: (510) 741-3941

Purpose for Submission:

New Device

Measurand:

IgM antibodies to Cardiolipin (CL) IgM antibodies to Beta-2-Glycoprotien I (ß2GPI)

Type of Test:

Semi-Quantitative multiplexed flow, bead-based immunoassay

Proprietary and Established Names:

BioPlex® 2200 APLS IgM kit BioPlex® 2200 APLS IgM Calibrator Set BioPlex® 2200 APLS IgM Control Set

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Regulatory Information:

1. Regulation section:
- 21 CFR §866.5660 Multiple autoantibodies immunological test system 21 CFR $862.1150 - Calibrator 21 CFR §862.1660 - Quality Control Material (assayed and unassayed)
1. Classification:

Class II (Assavs, Calibrator) Class 1 (Controls)

1. Product code:
  MID. System Test. Anti-Cardiolipin Immunological MSV. Antibodies. B2- Glycoprotein 1 (B2-GPI) JIX. Calibrator, Multi-Analyte Mixture JJX. Single (specified) Analyte Controls (Assayed and Unassayed)

4. Panel:

Immunology (82) (Assays) Chemistry (75) (Controls)

Intended Use:

l . Intended use(s):

The BioPlex 2200 APLS IgM kit is intended for use with the Bio-Rad BioPlex 2200 System.

The BioPlex 2200 Antiphospholipid Syndrome (APLS) IgM Calibrator Set is intended for the calibration of the corresponding BioPlex 2200 APLS IgM Reagent Pack.

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1. Indication(s) for use: Same as Intended Use
1. Special conditions for use statement(s): For prescription use only
1. Special instrument requirements: Bio-Rad BioPlex® 2200 Svstem

Device Description:

BioPlex® 2200 APLS IgM kit includes the following components:

o One (1) 10 mL vial of Bead Set containing two different populations of dyed beads coated with Cardiolipin (CL) and Beta-2-Glycoprotein I (B2GPI). an Internal Standard bead (ISB). a Serum Verification bead (SVB), and a Reagent Blank bead (RBB) in a MOPS (3-[N-Morpholino] propanesulfonic acid) buffer supplemented with glycerol and protein stabilizers (porcine), and ProClin 300 (< 0.3%), sodium benzoate (≤ 0.1%) and sodium azide (< 0.1%) as preservatives.
One (1) 5 mL vial of Conjugate containing phycoervthrin conjugated murine . monoclonal anti-human IgM and phycoerythrin conjugated murine monoclonal anti-human FXIII in MOPS (3-[N-Morpholino] propanesulfonic acid) buffer supplemented with protein stabilizers (bovine), and ProClin 300 (< 0.3%), sodium benzoate (≤ 0.1%) and sodium azide (< 0.1%) as preservatives.
One (1) 10 mL vial of Sample Diluent containing buffer with protein stabilizers . (bovine and murine), and ProClin 300 (< 0.3%). sodium benzoate (≤ 0.1%) and sodium azide (< 0.1%) as preservatives.

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Substantial Equivalence Information:

1. Predicate device name(s): HemosIL AcuStar anti-Cardiolipin IgM, K092181 HemosIL AcuStar anti-B2 Glycoprotein-1 1gM, K091556
  .

2. Comparison with predicate:

Similarities
Item	Device:	Predicate:	Predicate:
	BioPlex 2200 APLS IgM Kit	HemosIL Acustar anti-Cardiolipin IgM Kit	HemosIL Acustar anti-β2GPI IgM Kit
Intended Use	Semi-quantitative detection of IgM antibodies to Cardiolipin (CL) and Beta-2 Glycoprotein I (β2GPI) in human serum and plasma as an aid in the diagnosis of primary Antiphospholipid Syndrome (APS) and those secondary to systemic lupus erythematosus (SLE) or SLE-like disorders	Semi-quantitative measurement of anti-Cardiolipin (aCL) IgM antibodies in human citrated plasma or serum on the ACL AcuStar, as an aid in the diagnosis of thrombotic disorders related to primary and secondary Antiphospholipid Syndrome (APS)	Semi-quantitative measurement of Anti-B2 Glycoprotein-I (Anti-β2GPI) IgM antibodies in human plasma or serum on the ACL AcuStar, as an aid in the diagnosis of thrombotic disorders related to primary and secondary Antiphospholipid Syndrome (APS)
Sample Type	Serum or plasma (lithium heparin, sodium heparin, and sodium citrate)	Serum or citrated plasma	Serum or citrated plasma
Assay Type	Semi-quantitative	Same	Same
Analyte Detected	Human IgM antibodies to Cardiolipin	Same	Not Applicable
	Human IgM antibodies to β2GPI	Not Applicable	Same
Capture Antigen	Cardiolipin	Same	Not Applicable
	Human β2GPI	β2GPI	Same
Controls	Two Levels	Same	Same

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	Differences
Item	Device:BioPlex 2200 APLS IgM Kit	Predicate:HemosIL Acustar anti-Cardiolipin IgM Kit	Predicate:HemosIL Acustaranti-β2GPI IgM Kit
Assay Technology	Automated multiplex flowimmunoassay	Two-stepchemiluminescentimmunoassay	Two-stepchemiluminescentimmunoassay
ConjugateAntibody	Phycoerythrin conjugatedmurine monoclonal anti-human IgM	Isoluminol-labeledanti-human IgMantibody	Isoluminol-labeledanti-human IgMantibody
Signal Detection	Fluorescence	Chemiluminescent	Chemiluminescent
Solid Phase	Antigen-coated paramagneticmicrobead reagent	Antigen-coatedmagnetic particles	Antigen-coatedmagnetic particles
Calibrator(s)	4 calibrator levels (soldseparately)	Two calibrator levels(included in test kit)	Two calibrator levels(included in test kit)
Control	One negative and onepositive control (Soldseparately)	One low and onehigh control (Soldseparately)	One low and one highcontrols (Soldseparately)
Assay range	Anti-Cardiolipin:0.2 – 112 MPL-U/mL	1.0 – 15480 U/mL	Not Applicable
	Anti-Beta-2-Glycoprotein:0.2 – 112 U/mL	Not Applicable	1.1 – 16820 U/mL
Quantitation	Results are determined froma standard calibration curveutilizing a point-to-pointcurve fitting.	Assay utilizes astored Master 4Parameter LogisticCurve (4PLC) fitadjusted with two lotdependent calibratorlevels	Assay utilizes astored Master 4Parameter LogisticCurve (4PLC) fitadjusted with two lotdependent calibratorlevels
Instrumentation	Bio-Rad BioPlex 2200system	HemosIL AcuStar	HemosIL AcuStar

Standard/Guidance Document Referenced (if applicable):

CEN 13640:2002, Stability Testing of In Vitro Diagnostic Reagents

EP05-A2, Evaluation of Precision Performance of Quantitative Measurement Methods; Approved Guideline, Second Edition.

EP06-A, Evaluation of Linearity of Quantitative Measurement, Approved Guideline, Second Edition.

EP07-A2. Interference Testing in Clinical Chemistry, Approved Guideline, Second Edition EP09-A2IR, Method Comparison and Bias Estimation Using Patient Samples, Approved Guideline, Second Edition (Interim Revision) (used for matrix comparison).

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EP12-A2. User Protocol for Evaluation of Qualitative test Performance, Approved Guideline, Second Edition.

EP14-A2, Evaluation of Matrix Effects. Approved Guideline, Second Edition

EP15-A2, User Verification of Performance for Precision and Trueness, Approved Guideline, Second Edition.

EP17-A, Protocols for Determination of Limits of Detection and Limits of Quantification. Approved Guideline.

Test Principle:

L. Test Principle:

The BioPlex® 2200 APLS IgM kit uses multiplex flow immunoassay, a methodology similar to traditional EIA: however, this method permits simultaneous detection and identification of manv antibodies in a single tube. In the BioPlex APLS assavs. two different populations of dyed beads are coated with antigens. One bead population is coated with ß2-glycoprotein I and a second population is coated with both Cardiolipin and B2-glycoprotein 1. Three additional populations of fluorescent beads function as assay controls. The system combines an aliquot of patient sample diluent, and bead reagent into a reaction vessel and incubates the mixture at 37℃. After a wash cycle to remove unbound antibody. the secondary conjugate containing either phycoerythrin conjugated murine monoclonal antihuman IgM and phycoerythrin conjugated murine monoclonal anti-human FXIII antibody (a control) is added and the mixture is incubated at 37°C. Excess conjugate is removed in another wash cvcle and the beads are re-suspended in wash buffer. The bead mixture then passes through the detector. The identity of the dyed beads is determined by the fluorescence of the dyes, and the amount of antibody captured by the antigen is determined by the fluorescence of the attached phycoerythrin. Raw data are calculated in relative fluorescence intensity (RFI).

Three additional dyed beads. Internal Standard Bead (ISB), Serum Verification Bead (SVB), and a Reagent Blank Bead (RBB) are present in each reaction mixture to verify detector response, the addition of serum to the reaction vessel, and the absence of significant nonspecific binding in serum or plasma, respectively.

The anti-phospholipid assays are calibrated using a set of calibrators supplied separately by Bio-Rad Laboratories. Results are calculated for both of the antibodies and are compared against their own respective cut-off. For anti-B2-glycoprotein 1, the results are provided in units/ml. (U/mL). The results are similar except that the unit is MPL-U/mL for the IgM assay. The negative/positive assay cutoff for two analytes (anti-B2-glvcoprotein I and anti-Cardiolipin IgM) of the BioPlex® 2200 APLS IgM kit is 20 units.

Performance Characteristics (if/when applicable):

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

Precision testing of the BioPlex® 2200 APLS IgM kit on the BioPlex® 2200 instrument was performed in accordance with CLSI EP5-A2. Two serum and heparnized plasma panels consisting of samples spanning the measuring range were assayed in replicate twice daily over 20 davs (n=80) except for the mid negative samples were run over 10 days (n=40). One positive and one negative

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control were included. The data were analyzed for within-run, between-run. between-day, and total precision and the mean, standard deviation and percent coefficient of variation are summarized below:

Precision		Mean	Within Run		Between Run		Between Day		Total Precision
Sample	ટ	MPL-U/mL	SD	%CV	SD	%CV	SD	%CV	SD	%CV
Mid Negative	40	6.1	0.2	3.0%	0.1	1 .6%	0.1	I .5%	0.2	3.7%
Mid Negative	40	8.9	0.2	2.8%	0.1	1.6%	0.0	0.0%	0.2	3.2%
Mid Negative	40	9.1	0.3	2.8%	0.2	2.1%	0.0	0.0%	0.3	3.5%
Mid Negative	40	10.7	0.4	3.3%	0.1	1.1%	0.1	1.0%	0.4	3.6%
Negative NearCutoff	80	! 4.5	0.8	5.3%	0.5	3.6%	0.6	3.8%	1.1	7.4%
Negative NearCuloff	80	। રે રિ	0.9	6.0%	0.2	1.1%	0.1	0.4%	1.0	6.1%
Near Cut-Off	80	17.5	1 . 1	6.3%	0.2	1.0%	0.8	4.5%	1.4	7.9%
Near Cut-Off	80	19.6	1 .3	6.4%	0.4	2.1%	0.9	4.3%	1 .6	8.0%
Low Positive	80	22.6	1.0	4.5%	0.8	3.5%	1.0	4.6%	1.7	7.3%
Low Positive	80	22.7	1 .3	5.7%	0.8	3.4%	l . I	4.8%	1.9	8.2%
High Positive	80	73.6	5.5	7.5%	। .રે	2.1%	1.8	2.4%	6.0	8.1%
High Positive	80	79.1	3.7	4.7%	0.0	0.0%	2.1	2.7%	4.3	5.4%
Pos. Control	80	62.9	2.3	3.7%	1.9	3.0%	1.2	2.0%	3.2	5.2%

BioPlex® 2200 APLS IgM - Anti-Cardiolipin: Serum Samples

BioPlex® 2200 APLS IgM - Anti-Cardiolipin: Heparin Sample

Precision		MeanMPL-U/mL	Within Run		Between Run		Between Day		Total Precision
Sample	N		SD	%CV	SD	%CV	SD	%CV	SD	%CV
Mid Negative	40	6.8	0.2	2.7%	0.1	2.1%	0.0	0.7%	0.2	3.4%
Mid Negative	40	7.4	0.2	3.0%	0.1	1.3%	0.1	1.4%	0.3	3.6%
Mid Negative	40	10.4	0.3	3.1%	0.3	2.5%	0.0	0.0%	0.4	4.0%
Mid Negative	40	10.7	0.3	2.8%	0.0	0.0%	0.2	2.1%	0.4	3.5%
Negative NearCutoff	80	13.6	0.6	4.6%	0.2	1.6%	0.4	3.2%	0.8	5.8%
Negative NearCutoff	80	14.0	0.8	5.4%	0.2	1.3%	0.6	4.4%	1.0	7.1%
Near Cut-Off	80	18.2	1.0	5.3%	0.7	3.9%	0.2	1.1%	1.2	6.7%
Near Cut-Off	80	19.0	1.3	6.9%	0.3	1.3%	0.6	3.0%	1.4	7.6%
Low Positive	80	21.5	1.3	6.1%	0.4	1.8%	0.0	0.0%	1.4	6.3%
Low Positive	80	23.4	0.8	3.4%	0.7	2.8%	0.9	3.9%	1.4	5.9%
High Positive	80	72.7	3.2	4.4%	1.4	1.9%	2.1	2.9%	4.1	5.6%
High Positive	80	72.8	2.8	3.8%	2.2	3.1%	0.0	0.0%	3.6	4.9%

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PrecisionSample	N	MeanU/mL	Within Run		Between Run		Between Day		Total Precision
			SD	%CV	SD	%CV	SD	%CV	SD	%CV
Mid Negative	40	8.8	0.3	3.0%	0.1	0.7%	0.0	0.0%	0.3	3.1%
Mid Negative	40	9.0	0.3	3.1%	0.3	2.8%	0.0	0.0%	0.4	4.2%
Mid Negative	40	12.8	0.5	3.5%	0.4	2.9%	0.0	0.0%	0.6	4.6%
Mid Negative	40	15.4	0.6	4.1%	0.2	1.2%	0.3	1.7%	0.7	4.6%
Negative NearCutoff	80	17.4	0.6	3.3%	0.2	1.3%	0.1	0.7%	0.6	3.7%
Negative NearCutoff	80	17.7	0.7	3.8%	0.4	2.1%	0.4	2.1%	0.9	4.8%
Near Cut-Off	80	20.6	0.9	4.2%	0.0	0.0%	0.5	2.6%	1.0	4.9%
Near Cut-Off	80	21.4	0.8	3.6%	0.4	2.0%	0.6	2.7%	1.1	4.9%
Low Positive	80	23.5	0.7	3.1%	0.7	2.7%	0.7	2.9%	1.2	5.1%
Low Positive	80	23.5	0.9	3.9%	0.5	2.0%	0.9	3.7%	1.4	5.8%
High Positive	80	79.5	5.7	7.2%	2.5	3.2%	1.1	1.4%	6.3	8.0%
High Positive	80	85.5	3.9	4.6%	0.0	0.0%	2.0	2.4%	4.4	5.2%
Pos. Control	80	52.1	2.3	4.4%	1.7	3.2%	1.4	2.7%	3.2	6.0%

BioPlex® 2200 APLS IgM - Anti- B2GPI: Serum Samples

BioPlex® 2200 APLS IgM - Anti- B2GPI: Heparin Samples

PrecisionSample	N	MeanU/mL	Within Run		Between Run		Between Day		Total Precision
			SD	%CV	SD	%CV	SD	%CV	SD	%CV
Mid Negative	40	8.4	0.3	3.1%	0.2	1.8%	0.0	0.3%	0.3	3.7%
Mid Negative	40	8.6	0.3	3.3%	0.1	1.5%	0.1	1.0%	0.3	3.8%
Mid Negative	40	12.2	0.5	3.7%	0.3	2.7%	0.0	0.0%	0.6	4.6%
Mid Negative	40	13.1	0.4	2.8%	0.0	0.0%	0.2	1.5%	0.4	3.2%
Negative NearCutoff	80	16.8	0.5	3.0%	0.2	1.4%	0.4	2.5%	0.7	4.2%
Negative NearCutoff	80	17.0	0.7	3.9%	0.0	0.0%	0.5	2.7%	0.8	4.7%
Near Cut-Off	80	20.3	0.6	3.0%	0.6	2.9%	0.0	0.2%	0.9	4.2%
Near Cut-Off	80	20.9	0.8	3.9%	0.0	0.0%	0.5	2.3%	0.9	4.5%
Low Positive	80	23.3	0.9	3.9%	0.3	1.1%	0.0	0.0%	1.0	4.1%
Low Positive	80	24.0	0.6	2.4%	0.4	1.7%	0.6	2.6%	0.9	3.9%
High Positive	80	78.6	2.9	3.7%	2.3	2.9%	0.0	0.0%	3.7	4.7%
High Positive	80	79.0	3.5	4.5%	1.5	1.9%	1.4	1.8%	4.1	5.2%

CLSI EP15-A2 Reproducibility

Precision and reproducibility was also evaluated in accordance with CLSI EP15-A2 guideline "User Verification of Performance for Precision and Trueness, Vol 25, No 17".

A different serum panel consisting of samples spanning the measuring range were assayed in 4 replicates per run, one run per day over 5 days (n=20). One positive and one negative control were included. The data were analyzed for within-run, between run (day), and total precision and the mean U/mL, standard deviation and percent coefficient of variation are summarized below:

{8}------------------------------------------------

Panel Member	N	Mean	Within Run		Between Run		Total
			SD	%CV	SD	%CV	SD	%CV
Negative 1	20	12.2	0.42	3.5 %	0.55	4.5 %	0.69	5.7 %
Negative 2	20	14.7	0.87	5.9 %	1.01	6.9 %	1.34	9.1 %
Near Cutoff 1	20	17.8	1.43	8.0 %	1.26	7.1 %	1.91	10.7 %
Near Cutoff 2	20	19.6	0.90	4.6 %	0.77	3.9 %	1.18	6.0 %
Low positive 1	20	35.0	2.84	8.1 %	1.02	2.9 %	3.01	8.6 %
Low positive 2	20	41.1	2.50	6.1 %	1.45	3.5 %	2.89	7.0 %
High Positive 1	20	64.6	3.43	5.3 %	2.30	3.6 %	4.13	6.4 %
High Positive 2	20	73.9	5.71	7.7 %	0.00	0.0 %	5.71	7.7 %
Negative Control	20	<0.2	N/A
Positive Control	20	43.7	2.67	6.1 %	0.00	0.0 %	2.67	6.1 %

BioPlex® 2200 APLS IgM - Anti- Cardiolipin: Serum Samples

BioPlex® 2200 APLS IgM - Anti- ß2GPI: Serum Samples

			Within run			Between Run	Total
Panel Member	N	Mean	SD	%CV	SD	%CV	SD	%CV
Negative 1	20	12.2	0.58	4.7 %	0.73	ર છે જે	0.93	7.6 %
Negative 2	20	17.5	0-85	4.9 %	0.62	3.6 %	1.05	6.0 %
Near Cutoff I	20	18.5	l . I l	6.0 %	1.07	5.8 %	1.54	8.3 %
Near Cutoff 2	20	22.2	1.73	7.8 %	1.44	6.5 %	2.25	10.1 %
Low positive I	20	52.8	4.36	8.3 %	0.00	0.0 %	4.36	8.3 %
Low positive 2	20	53.4	3.16	5.9 %	1.62	3.0 %	3.55	6.7 %
High Positive 1	20	92.9	6.74	7.3 %	0.00	0.0 %	6.74	7.3 %
High Positive 2	20	96.2	5.30	5.5 %	1.52	1.6 %	ર .5 l	5.7 %
Negative Control	20	<0.2				N/A
Positive Control	20	55.6	3.24 .	5.8 %	0.00	0.0 %	3.24	5.8 %

Lot-to-lot Reproducibility

The lot-to-lot reproducibility study was conducted to evaluate the variation among the lots of the reagent kit. Serum samples spanning the assay range were tested with three reagent lots on three BioPlex 2200 instruments in replicates of 10 for two runs.

Each lot mean (MPL-U/mL for aCL IgM and U /mL for aß2GPI IgM) was calculated using data 60 points for each patient serum sample (ten replicates, three instruments, two runs per instrument). The lot to lot grand mean (U/mL), standard deviation and %CV were calculated for each of the samples.

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	Mean	#		Within Run		BetweenRun		BetweenInstrument		Between Lot		Total
Accession	MPLU/mL	RunsperDay	#Days	SD	%CV	SD	%CV	SD	%CV	SD	%CV	SD	%CV
APSQSM03	15.0	1	2	0.42	2.8	0.42	2.8	0.00	0.0	0.59	3.9	0.84	5.6
APSQSM04	14.6	1	2	0.46	3.1	0.59	4.0	0.00	0.0	1.55	10.6	1.72	11.8
APSQSM06	23.6	1	2	0.73	3.1	0.78	3.3	0.00	0.0	1.31	5.6	1.69	7.2
APSQSM07	26.2	1	2	0.86	3.3	0.60	2.3	0.00	0.0	1.50	5.7	1.83	7.0
APSQSM14	91.9	1	2	3.40	3.7	2.63	2.9	0.00	0.0	2.73	3.0	5.09	5.5

Anti- Cardiolipin IgM: Lot-to-Lot Reproducibility

Anti- ß2GPI lgM: Lot-to-Lot Reproducibility

		#RunsperDay	#Days		Within Run		BetweenRun		BetweenInstrument		Between Lot		Total
Accession	MeanU/mL			SD	%CV	SD	%CV	SD	%CV	SD	%CV	SD	%CV
APSQSM03	16.5	1	2	0.51	3.1	0.38	2.3	0.00	0.0	0.92	5.6	1.12	6.8
APSQSM04	18.6	1	2	0.65	3.5	0.74	4.0	0.00	0.0	2.44	13.1	2.63	14.1
APSQSM06	22.9	1	2	0.72	3.2	0.56	2.5	0.00	0.0	1.40	6.1	1.67	7.3
APSQSM07	26.6	1	2	0.92	3.5	0.69	2.6	0.00	0.0	2.11	7.9	2.41	9.1
APSQSM14	98.0	1	2	3.88	4.0	3.03	3.1	0.00	0.0	10.01	10.2	11.16	11.4

b. Linearity/assay reportable range:

Six aCL and aB2GP1 IgM positive patient samples were tested to demonstrate linearity. These samples were diluted with immunodepleted serum according to CLSI EP06-A. Each sample and dilution was evaluated in replicates of four using one APLS IgM lot on one instrument. Linear and polynomial regression analysis of APLS IgM recovery vs. sample dilution was performed to determine if the dilution curves exhibit statistically significant non-linear regression based on the CLSI guideline EP06-A.

The regression parameters (slope, intercept and r2) of the observed values vs. predicted values are show below.

APLS IgMAssays	Sample	Conc.	Slope	Intercept	r²	Dilution range
Anti-CardiolipinIgM(MPL-U/mL)	1	49.5	1.0000	-0.0010	0.9931	0.6 - 49.5
	2	57.4	1.0005	-0.0117	0.9914	0.1 - 57.4
	3	54.1	0.9992	0.0184	0.9915	0.1 - 54.1
	4	93.9	0.9999	0.0174	0.9954	0.1 - 93.9
	5	92.3	0.9995	0.0203	0.9985	0.1 - 92.3
	6	100.0	0.9998	0.0226	0.9925	0.6 - 100.0
Anti-β2GPIIgM(U/mL)	1	54.2	0.9996	0.0099	0.9942	0.6 - 54.2
	2	49.9	1.0009	-0.0319	0.9933	0.1 - 49.9
	3	53.1	0.9996	0.0089	0.9952	0.1 - 53.1
	4	92.5	0.9996	0.0167	0.9978	0.6 - 92.5
	5	111.1	0.9998	0.0258	0.9980	0.1 - 111.1

{10}------------------------------------------------

			A more annual a c100.3	1.0003------------------------------------------------------------------------------------------------------------------------------------------------------------------------------	-0.0275	0.9985	0.2 - 100.3
--	--	--	----------------------------	------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------	---------	--------	-------------

BioPlex 2200 APLS IgM Assay	Assay reportable range
Anti-Cardiolipin IgM	0.2 to 112 MPL-U/mL
Anti-β2GPI IgM	0.2 to 112 U/mL

Over-Range (OR) results mav be generated for values greater than the reportable measuring range and results are reported as > 112 MPL-U/mL for aCL and > 112 U/mL for aß2GPI IgM.

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

Traceability:

There is no international or certified reference material available for anti-CL and anti-B2GPI IgM. The calibrators are assigned relative arbitrary units (MPL-U/mL for aCL and U/mL for aß2GPI).

Value Assignment:

The calibrators are manufactured independently from the controls, and are stabilized with <0.3% ProClin® 300. <0.1% sodium benzoate, and <0.1% sodium azide. Calibrator assignment is established for matched lots of BioPlex® 2200 APLS IgM kit and calibrators using a master set of calibrators as reference and replicate analyses on multiple BioPlex® 2200 instruments. The BioPlex® 2200 APLS IgM Reagent Kit is calibrated using a set of four (4) distinct calibrators for anti-Cardiolipin and anti-B2GPI IgM, which are used to establish points of reference for determining the presence of anti-Cardiolipin or anti-B2GPI IgM in human specimens.

The negative control has been tested to give results with values below the cut-off for each assay. The positive control is prepared by blending human disease state serum with negative scrum matrix and is manufactured to give results with values above the assay cut-off. The positive controls are manufactured to give positive results, with values above the cut-off for each specific analyte. The negative control is manufactured to give negative results, with values below the cut-off for each specific analyte.

Stability:

Stability studies have been performed to support the following claims:

Calibrator and Control:

BioPlex® 2200 APLS IgM Control and Calibrator Sets: Calibrator Open Vial Stability (2 to 8°C). 30 days from first opening: Control Open Vial Stability (2 to 8ºC). 60 days from first opening: Onboard Calibration Curve Stability. 30 days: Calibrators and Controls Real Time Stability (2 to 8°C), 24 months: labeled as until expiration date; Calibrators and Controls Accelerated Stability (2 to 8℃),

{11}------------------------------------------------

2.5 years predicted. Calibrators and Controls freeze-thaw (-20℃ or -70℃), 5freeze thaw cycles.

Kit Stability:

BioPlex® 2200 APLS IgM Kit: Real Time (unopened) Kit Stability, 24 months or until the date of expiration when stored unopened on the instrument or at 2 to 8°C: The Open kit claim is 60 days.

Sample Stability:

Sample stability studies were also performed: Sample stability fresh (2 to 8°C), 7 days: Sample stability frozen (-20 or -70°C). 24 months: Sample Freeze-thaw (-20 or -70℃), up to 3-freeze thaw cycles acceptable.

d. Detection limit:
The results of LoQ, LoD and LoB are summarized in the table below

BioPlex® 2200 APLS IgM Assay	LoQ	LoD	LoB
Anti-Cardiolipin IgM (MPL-U/mL)	0.2	0.13	0.0974
Anti-β2GPI IgM (U/mL)	0.2	0.13	0.1044

Analytical specificity: હ.
An interfering substances study was conducted to evaluate the potential interference of specific endogenous and exogenous substances with the BioPlex® 2200 APLS IgM kit according to CLSI EP7-A2. No interference was observed with any of the substances tested. The substances and the maximum levels tested are shown in the table below:

Substance	Concentration
Hemoglobin	≤500 mg/dL
Bilirubin (unconjugated)	≤20 mg/dL
Bilirubin (conjugated)	≤30 mg/dL
Triglycerides	≤3300 mg/dL
Total Protein	≤12 g/dL
Cholesterol	≤500 mg/dL
Red Blood Cells	≤0.4% (v/v)
Gamma-Globulin	≤6 g/dL
Beta-Carotene	≤0.6 mg/dL
Ascorbic Acid	≤3 mg/dL
Heparin Lithium	≤8000 units/dL
Heparin Sodium	≤8000 units/dL
Sodium Citrate	< 1000 mg/dL

Cross-Reactivity:

A cross-reactivity study was performed to determine if samples from individuals with various disease states and other potentially interfering factors interfere with

{12}------------------------------------------------

test results from the BioPlex® 2200 APLS IgM kit. Samples from individuals with known disease states for potential cross reactivity listed in the table below were evaluated with the BioPlex® 2200 APLS IgM kit. The table below shows the number (N) of samples containing potential cross reactants as disease state evaluated by the BioPlex® APLS IgM kit. The cross reactivity was obtained as the positivity rate from the ratio of the number of samples scored positive by the BioPlex® APLS IgM assays to the total number of cross reactant samples evaluated.

		BioPlex 2200 APLS IgM
Cross ReactiveDisease State	N	Anti-CL IgM		Anti-β2GPIIgM
		Pos (+)	% Positivity	Pos (+)	% Positivity
Systemic LupusErythematosus	34	2	5.9%	2	5.9%
Scleroderma	20	3	15.0%	3	15.0%
Sjogrens	22	0	0%	0	0%
Crohn's Disease	21	2	9.5%	2	9.5%
Ulcerative Colitis	20	1	5.0%	1	5.0%
Rheumatoid Arthritis	12	1	8.3%	1	8.3%
Syphilis	15	0	0%	0	0%

High dose hook effect: Not Applicable

f. Assay cut-off:

The cutoff value and assignment of the calibrators are determined by performing concordance testing and Receiver Operator Characteristic (ROC) analysis using the clinical diagnosis as the standard. The study to determine the APLS IgM assay cutoff is comprised of two sample groups - one clinical cohort has 103 samples from patients diagnosed as primary or secondary APS and 123 from non-APS or other rheumatic disease control donors. It was later confirmed by testing 208 samples from apparently healthy donors.

The cut-off was established to achieve a clinical specificity of 99% while accepting the resultant clinical sensitivity. The criteria for choosing a cutoff at the

{13}------------------------------------------------

9911 percentile of a normal healthy population is derived from the "International Consensus Statement on an Update of the Classification Criteria for Definite Antiphospholipid Syndrome (APS)". Journal of Thrombosis and Haemostasis (2006)4, 295-306.

A cutoff of 20.0 MPL-U/mL for anti-CL IgM and 20 U/mL for anti- B2GPI IgM was established by optimizing for clinical specificity.

2. Comparison studies:

Method comparison with predicate device: તા.
The performance of the BioPlex® 2200 APLS IgM kit was evaluated including 199 patients diagnosed with primary or secondary APS, 346 patients with other rheumatic or non-APS disease. Results in the measuring range and 10% of diluted total samples of both the new and the predicate immunoassays are compared. Results are summarized in the tables below:

		Predicate IgM Kit(aCL: 1.0 - 15480 U/mL; aβ2GP1:1.1 - 16820 U/mL)
Diagnosed Primary and Secondary APS Patientsand Non-APS Disease Patients		Positive	Negative	Total
aCL IgM(0.2 -112 MPL-U/mL)	Positive	77	1	78
	Negative	28	403	431
	Total	105	404	509
aβ2GPI IgM(0.2 - 112 U/mL)	Positive	79	15	94
	Negative	4	233	237
	Total	83	248	331

BioPlay

aCL IgM Positive Agreement (95% CI) = 73.3% (77/105) (64.2 - 80.9%) aCL IgM Negative Agreement (95% CI) = 99.8% (403/404) (98.6 - 100%) aCL IgM Total Agreement (95% CI) = 94.3% (480/509) (91.9 - 96.0%) aß2GPI IgM Positive Agreement (95% Cl) = 95.2% (79/83) (88.3 - 98.1%) aß2GPI IgM Negative Agreement (95% CI) = 94.0% (233/248) (90.3 - 96.3%) aß2GPI IgM Total Agreement (95% Cl) = 94.3% (312/331) (91.2 - 96.3%)

b. Matrix comparison:

Testing for matrix effects was conducted using 38 matched sets of serum. heparin, and citrate plasma samples drawn from the same donor in accordance with CLSI EP9-A2. The samples were spiked with aCL IgM or aß2GPI IgM positive sera as necessary in order to assemble a panel of samples to cover the measuring range of the assay. All samples were evaluated in replicates of two. Plasma values were compared to matched serum values. Anticoagulants were

{14}------------------------------------------------

considered non-interfering if the linear regression of aCL IgM or aß2GPI IgM values from matched serum versus plasma samples has a slope of 1.00 ± 0.2, a yintercept of 0.0 ± 6.0 and a correlation coefficient (r) between 0.980 and 1.000. The regression correlation parameters for the slopes, intercepts and correlation coefficient (r) are summarized below.

Comparison	N	BioPlex® APLSAssay	Slope(95% CI)	Intercept(95% CI)	r
LithiumHeparin vs.Serum	38	aCL IgM	1.0106(0.9808, 1.0405)	-0.3053(-2.0421, 1.4315)	0.9962
		aβ2GPI IgM	0.9991(0.9682, 1.0301)	0.3054(-1.5299, 2.1405)	0.9958
SodiumHeparin vs.Serum	38	aCL IgM	1.0369(0.9786, 1.0953)	-0.6412(-4.0349, 2.7525)	0.9864
		aβ2GPI IgM	1.0271(0.9750, 1.0779)	-0.2328(-3.3212, 2.8555)	0.9889
SodiumCitrate vs.Serum	38	aCL IgM	1.0494(1.0190, 1.0779)	-0.9676(-2.7381, 0.8029)	0.9963
		aβ2GPI IgM	1.0354(1.0082, 1.0627)	-0.4738(-2.0890, 1.1414)	0.9970

3. Clinical studies:

a. Clinical Sensitivity and specificity:

The clinical studies involved testing 545 specimens including 199 diagnosed primary or secondary APS patients and 346 non-APS disease control patients. The BioPlex® 2200 APLS IgM sensitivity and specificity are shown below:

	Clinical Sensitivity and Specificity	Clinical Diagnosis
		Diagnosed with Primary or Secondary APS	Non-APS Disease Controls	Total
aCL IgM	Positive	67	11	78
	Negative	132	335	467
	Total	199	346	545
aβ2GPI IgM	Positive	80	14	94
	Negative	119	332	451
	Total	199	346	545

BioPlex 2200 APLS IgM
-----------------------	--

aCL IgM Sensitivity (95% CI) = 33.7% (67/199) (27.5 - 40.5%) aCL IgM Specificity (95% Cl) = 96.8% (335/346) (94.4 - 98.2%)

{15}------------------------------------------------

aB2GP1 IgM Sensitivity (95% CI) = 40.2% (80/199) (33.6-47.1%) aB2GPI IgM Specificity (95% CI) = 96.0% (332/346) (93.3 - 97.6%)

Disease Category	NumberEnrolled	aCL IgM	aβ2GPI IgM
Apparently Healthy Subject	300	7	3
Primary APS (PAPS)	123	43	50
Secondary APS (SAPS)	76	24	30
Syphilis	15	0	0
Systemic LupusErythematosus	101	4	5
Rheumatoid Arthritis	90	3	4
Crohn's Disease	21	1	1
CREST	3	0	0
Fibromyalgia	19	0	0
Gout	14	0	0
Inflammatory Arthritis	4	0	0
Osteoarthritis	11	1	1
Scleroderma	24	1	3
Sjogrens	21	1	0
Ulcerative Colitis	18	0	0
Wegeners Granulomatosis	5	0	0

The number of samples positive of the BioPlex APLS IgM assay in each of disease category are shown below

c. Other clinical supportive data (when a. and b. are not applicable):
Refer to Method Comparison
1. Clinical cut-off: See Assay Cutoff
1. Expected values/Reference range:

Three hundred samples from apparently healthy donors including 132 males ranging in age from 7 to 85 and 168 females ranging in age from 14 to 83 were tested with BioPlex® 2200 APLS IgM kit. The number of positive and mean value of the BioPlex® APLS IgM results is shown below.

Assay	N (%Positive)	Mean	99th Percentile
aCL IgM	7 (2.3%)	3.0 MPL-U/mL	27.9 MPL-U/mL
aβ2GPI IgM	3 (1.0%)	2.5 U/mL	19.4 U/mL

Each laboratory should establish its own reference range pertinent to their specific patient populations.

{16}------------------------------------------------

Instrument Name:

。

The BioPlex 2200 System, software version 4.0 has been cleared in K103834.

and the state of the state of the states of the states of the states of the states of the states of the states of the states of the states of the states of the states of the

1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 - 1999 -

、

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{17}------------------------------------------------

DEPARTMENT OF HEALTH & HUMAN SERVICES

Image /page/17/Picture/1 description: The image shows the logo for the U.S. Department of Health & Human Services. The logo consists of a stylized eagle with its wings spread, and the words "DEPARTMENT OF HEALTH & HUMAN SERVICES USA" are arranged in a circular pattern around the eagle. The logo is black and white.

Public Health Service

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

October 21, 2013

BIO-RAD LABORATORIES DR. JUANG WANG REGULATORY AFFAIRS REPRESENTATIVE 5500 E. 2ND STREET BENICIA CA 94510

Re: K130528

Trade/Device Name: BioPlex 2200 APLS IgM kit, BioPlex 2200 APLS IgM Calibrator Set, BioPlex 2200 APLS IgM Control Set Regulation Number: 21 CFR 866.5660 Regulation Name: Multiple autoantibodies immunological test system Regulatory Class: Il Product Code: MID, MSV, JIX, JJX Dated: October 01, 2013

Received: October 10, 2013

Dear Dr. Wang:

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

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

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

{18}------------------------------------------------

Page 2-Dr. Juang Wang

electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR 1000-1050.

If you desire specific advice for your device on our labeling regulations (21 CFR Parts 801 and 809), please contact the Division of Small Manufacturers, International and Consumer Assistance at its toll-free number (800) 638 2041 or (301) 796-7100 or at its Internet address http://www.fda.gov/MedicalDevices/ResourcesforYou/Industry/default.htm. Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21CFR Part 807.97). For questions regarding the reporting of adverse events under the MDR regulation (21 CFR Part 803), please go to

http://www.fda.gov/MedicalDevices/Safety/ReportalProblem/default.htm for the CDRH's Office of Surveillance and Biometrics/Division of Postmarket Surveillance.

You may obtain other general information on your responsibilities under the Act from the Division of Small Manufacturers, International and Consumer Assistance at its toll-free number (800) 638-2041 or (301) 796-7100 or at its Internet address http://www.fda.gov/MedicalDevices/ResourcesforYou/Industry/default.htm.

Sincerely yours,

Image /page/18/Picture/6 description: The image shows the text "Maria M. Chan - S". The text is written in a bold, sans-serif font. The letters "M.D.C" are stylized with a concentric line pattern. The rest of the text is in a plain, bold font.

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

Enclosure

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DEPARTMENT OF HEALTH AND HUMAN SERVICES Food and Drug Administration

Indications for Use

510(k) Number (if known) K130528

Device Name

BioPlex® 2200 Antiphospholipid Syndrome (APLS) IgM Kit

Indications for Use (Describe)

The BioPlex® 2200 Antiphospholipid Syndrome (APLS) IgM kit is a multiplex Aow immunossay intended for the semi-quantitative detection of IgM antibodies to Cardiolipin (CL) and Beta-2 Glycoprotein 1 (B2GPI) in human serum and plasma (Ithium heparin, sodium heparin. and sodium citrate). In conjunction with other clinical findings, the test system is used as an aid in the diagnosis of primary Antiphospholipid Syndrome (APS) and those secondary to systemic lupus crythematosus (SLE) or SLE-like disorders.

The BioPlex 2200 APLS IgM kit is intended for use with the Bio-Rad BioPlex 2200 System

Type of Use (Select one or both, as applicable)

X Prescription Use (Part 21 CFR 801 Subpart D)

Over-The-Counter Use (21 CFR 801 Subpart C)

PLEASE DO NOT WRITE BELOW THIS LINE - CONTINUE ON A SEPARATE PAGE IF NEEDED.

FOR FDA USE ONLY

Concurrence of Center for Devices and Radiological Health (CDRH) (Signature)

Image /page/19/Picture/14 description: The image shows the name "Maria M. Chan -S" in a bold, sans-serif font. The letters are black and have a slightly rough or textured appearance. The name is written in a single line, with a space between each word and initial. The letters "FC" are stylized with a box around them.

Form Approved: OMB No. 0910-0120 Expiration Date: December 31, 2013 See PRA Statement on last page.

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DEPARTMENT OF HEALTH AND HUMAN SERVICES Food and Drug Administration

Indications for Use

510(k) Number (if known) K130528

Device Name

BioPlex® 2200 Antiphospholipid Syndrome (API.S) IgM Control Set

Indications for Use (Describe)

The BioPlex 2200 Antiphospholipid Syndrome (APLS) IgM Control Set is intended for use as an assayed quality control to monitor the overall performance of the BioPlex 2200 Instrument and the corresponding BioMex 19th the clinical laboratory. The performance of the BioPlex 2200 APLS IgM Control Set has not been established with any other Antiphospholipid assay .

Type of Use (Select one or both, as applicable)

Prescription Use (Part 21 CFR 801 Subpart D)

[] Over-The-Counter Use (21 CFR 801 Subpart C)

PLEASE DO NOT WRITE BELOW THIS LINE - CONTINUE ON A SEPARATE PAGE IF NEEDED.

FOR FDA USE ONLY

Concurrence of Center for Devices and Radiological Health (CDRH) (Signature)

Image /page/20/Picture/13 description: The image shows the text "Maria M. FC Chan -S" in a bold, sans-serif font. The letters are black and have a slightly rough or textured appearance. The letters "FC" are stylized with a geometric pattern, making them stand out from the rest of the text.

Form Approved: OMB No. 0910-0120 Expiration Date: December 31, 2013 See PRA Statement on last page.

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DEPARTMENT OF HEALTH AND HUMAN SERVICES Food and Drug Administration

Indications for Use

510(k) Number (if known) K 130528

Device Name

BioPlex® 2200 Antiphospholipid Syndrome (API.S) IgM Calibrator Set

Indications for Use (Describe)

The BioPlex 2200 APLS IgM Calibrator Set is intended for the corresponding BioPlex 2200 APLS IgM Reagent Pack.

Type of Use (Select one or both, as applicable)

Prescription Use (Part 21 CFR 801 Subpart D)

Over-The-Counter Use (21 CFR 801 Subpart C)

PLEASE DO NOT WRITE BELOW THIS LINE -- CONTINUE ON A SEPARATE PAGE IF NEEDED.

FOR FDA USE ONLY

Concurrence of Center for Devices and Radiological Health (CDRH) (Signature)

Image /page/21/Picture/13 description: The image shows the name "Maria M. FC Chan -S" in a stylized font. The letters are large and bold, with a slightly rough, hand-drawn appearance. The name is arranged horizontally, with the first name "Maria" followed by the middle initial "M.", then "FC Chan", and finally "-S".

Form Approved: OMB No. 0910-0120 Expiration Date: December 31, 2013 See PRA Statement on last page.

Regulation Number and Section

§ 866.5660 Multiple autoantibodies immunological test system.

(a)
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).(b)
Classification. Class II (performance standards).