IBL C3D-CIC EIA TEST
Device Facts
| Record ID | K032796 |
|---|---|
| Device Name | IBL C3D-CIC EIA TEST |
| Applicant | Ibl GmbH |
| Product Code | CZW · Immunology |
| Decision Date | Jan 13, 2004 |
| Decision | SESE |
| Submission Type | Traditional |
| Regulation | 21 CFR 866.5240 |
| Device Class | Class 2 |
Indications for Use
The IBL C3d-CIC test is a semi-quantitative enzyme immunoassay for the in vitro diagnostic detection of circulating immune complexes that bind C3d in human serum. The measurement is performed as an aid in the diagnosis of various autoimmune and other CIC related diseases. Levels of these complexes are one indicator in a multifactorial diagnostic regime
Device Story
The IBL C3d-CIC EIA test is an in vitro diagnostic enzyme immunoassay used in clinical laboratory settings. It detects circulating immune complexes (CIC) that bind C3d in human serum samples. The assay functions as a semi-quantitative tool to assist clinicians in diagnosing autoimmune and CIC-related conditions. Results are intended to be used as one component of a multi-factorial diagnostic evaluation. The device provides laboratory professionals with a measured level of immune complexes, which supports clinical decision-making regarding the presence or progression of autoimmune disease.
Clinical Evidence
No clinical data provided; bench testing only.
Technological Characteristics
Sandwich ELISA; monoclonal anti-C3d antibody coated microplates; horseradish peroxidase-labeled anti-human IgG detection; TMB substrate; optical density measurement at 450 nm. Semi-quantitative; results expressed in µg/ml. Serum matrix only.
Indications for Use
Indicated for the semi-quantitative detection of C3d-binding circulating immune complexes (CIC) in human serum as an aid in the diagnosis of autoimmune and other CIC-related diseases.
Regulatory Classification
Identification
A complement components immunological test system is a device that consists of the reagents used to measure by immunochemical techniques complement components C1q , C1r , C1s , C2 , C3 , C4 , C5 , C6 , C7 , C8 , and C9 , in serum, other body fluids, and tissues. Complement is a group of serum proteins which destroy infectious agents. Measurements of these proteins aids in the diagnosis of immunologic disorders, especially those associated with deficiencies of complement components.
Related Devices
- K032254 — IBL C1Q-CIC CIA TEST · Ibl GmbH · Aug 29, 2003
- K031563 — BINDAZYME CLQ CIC EIA KIT · The Binding Site, Ltd. · Sep 10, 2003
- K012576 — CIC-C1Q EIA · American Laboratory Products Co., Ltd. · Jan 29, 2002
Submission Summary (Full Text)
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510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION
DECISION SUMMARY
DEVICE ONLY
A. 510(k) Number:
k032796
B. Analyte:
Complement component C3d – human IgG circulating immune complexes
C. Type of Test:
Semi-quantitative enzyme immunoassay
D. Applicant:
Lehnus and Associates for Immuno Biological Laboratories – Hamburg
E. Proprietary and Established Names:
IBL C3d-CIC Test Kit; complement C3 antigen, antiserum, control
F. Regulatory Information:
1. Regulation section:
21 CFR 866.5240 Complement components immunological test system
2. Classification:
Class II
3. Product Code:
CZW
4. Panel:
Immunology 82
G. Intended Use:
The IBL C3d-CIC test is a semi-quantitative enzyme immunoassay for the in vitro diagnostic detection of circulating immune complexes that bind C3d in human serum. The measurement is performed as an aid in the diagnosis of various autoimmune and other CIC related diseases. Levels of these complexes are one indicator in a multifactorial diagnostic regime
1. Indication(s) for use:
Aid in the diagnosis of various autoimmune and other CIC related diseases
2. Special condition for use statement(s):
None
3. Special instrument Requirements:
None
H. Device Description:
The assay is a sandwich solid phase enzyme immunoassay (ELISA) utilizing coated microplates. Microplates coated with antibody recognizing the C3d portion of C3d-
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circulating immune complexes capture complexes from patient serum samples, calibrators, or controls. Patient serum samples and controls are diluted 1:5 with sample diluent. Horseradish peroxidase labeled anti-human IgG is added after initial incubation and washing of the specimen in the coated microplate wells. After a second incubation and washing, enzyme substrate (tetramethylbenzidine; TMB) is added to develop color when enzyme is captured onto wells of the plate. The color reaction is stopped after 15 minutes at room temperature (18-24°C) by addition of stop solution. The amount of color produced in each well is measured in a microtiter plate reader capable of reading optical density at 450 nm. The amount of color developed by standards of previously determined concentration is proportional to the concentration of calibrators. The concentration of samples or controls is calculated from the linear relationship of absorbance with concentration of the calibrators. Calibrators of 2 different arbitrary concentrations are utilized to derive sample concentration, expressed as µg/ml. Positive and negative controls are also included to monitor the satisfactory performance of the assay.
I. Substantial Equivalence Information:
1. Predicate device name(s): Scimedx Corp. C3d ScanLisa
2. Predicate K number(s): K861362
3. Comparison with predicate:
| Similarities | | |
| --- | --- | --- |
| Item | Device | Predicate |
| Assay technology | Enzyme immunoassay | Enzyme immunoassay |
| Analyte capture mechanism | Monoclonal anti-C3d antibody coated microplates | Monoclonal anti-C3d coated microplates |
| Calibration material | Heat aggregated human IgG | Heat aggregated human IgG |
| Differences | | |
| Item | Device | Predicate |
| Calibrators | 2 of different concentration | 3 of different concentration |
| Equivocal zone | 16-24 μg/ml | No equivocal zone |
| Intended Use | A semi-quantitative enzyme immunoassay for the in vitro diagnostic detection of circulating immune complexes that bind C3d in human serum. The measurement is performed as an aid in the diagnosis of various autoimmune and other CIC related diseases. Levels of these complexes are one indicator in a multi-factorial diagnostic regime | Not stated in labeling |
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J. Standard/Guidance Document Referenced (if applicable):
None
K. Test Principle:
Enzyme immunoassay utilizing antibody to complement component C3d coated to microwells to capture complexes of human IgG containing complement component C3d. Enzyme-labeled antibody to human IgG recognizes captured complexes absorbed on microwells.
L. Performance Characteristics (if/when applicable):
1. Analytical performance:
a. Precision/Reproducibility:
To assess inter-day reproducibility, 5 samples of varying concentration were tested on 8 days. The samples included 3 with positive concentrations, 1 with an equivocal concentration, and 1 with a negative concentration. To assess intra-assay imprecision, 5 samples similar to the inter-day description of imprecision were tested in 12 replicates. For each sample on each day the mean, standard deviation, and %coefficient of variation (CV) were calculated.
| Precision | Range (μg/ml) | CV(%) |
| --- | --- | --- |
| Intra-Assay | 4.0 – 83.6 | 8.1 – 3.2 |
| Inter-Assay | 4.0 – 83.0 | 12.3 – 2.5 |
b. Linearity/assay reportable range:
A high positive sample was serially diluted from 1:0.625 to 1:20 with diluent buffer. The observed concentration was determined from a standard curve developed by linear regression analysis of the optical density and the concentration of the blank, standard, positive control and negative control. From this analysis, an expected concentration for the serial dilutions of the sample was determined. The observed concentration and expected concentration were compared by regression analysis to determine if the slope was equivalent with 1.0. The slope of the regression line of observed vs. expected C3d CIC concentration was 1.14 (95% confidence interval 0.87 to 1.41). The correlation coefficient was 0.986. The slope is equivalent with 1.0 (i.e. the confidence interval contains 1.0). Therefore, the dilution analysis indicates linearity of observed C3d concentration with dilution of the sample up to 1:20.
c. Traceability (controls, calibrators, or method):
Values for standards and controls were determined from standards in the predicate assay. Two approximate values were chosen, 100
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$\mu \mathrm{g} / \mathrm{ml}$ and $10~\mu \mathrm{g / ml}$ . Values for controls are assigned an acceptable range based on the mean $\pm 2$ standard deviations.
d. Detection limit:
Sample diluent buffer was tested in 26 replicates on a single plate. The concentration was calculated using the standard curve. The mean value was $1.9~\mu \mathrm{g / ml}\pm 1$ standard deviation (SD) $0.13~\mu \mathrm{g / ml}$ . The limit of detection was defined as the mean concentration plus 3 SDs. The limit of detection is $2.2~\mu \mathrm{g / ml}$ .
e. Analytical specificity:
Four samples of different concentrations were obtained and supplemented with 4 concentrations of 3 different interfering substances. The 4 samples ranged in concentration from 10 to 37 $\mu \mathrm{g} / \mathrm{ml}$ . Bilirubin at concentrations from $1\mathrm{mg / dL}$ to $250\mathrm{mg / dL}$ caused less than $\pm 25\%$ interference at the concentrations tested (10 - 37 $\mu \mathrm{g} / \mathrm{ml}$ ). Hemoglobin at concentrations from $100\mathrm{mg / dL}$ to $1000\mathrm{mg / dL}$ caused less than $\pm 25\%$ interference at the concentrations tested. Finally, EDTA at concentrations ranging from 1 to $50\mathrm{mM}$ caused less than $\pm 15\%$ interference at the concentrations tested.
Monoclonal anti-C3d utilized to capture complement component C3d in the assay does not cross-react with human IgG but due to the structure of C3d will cross-react with C3 and C4 and not C3b.
f. Assay cut-off:
A gray zone was initially suggested by utilizing a normal population (using 3 and 5 standard deviations above the mean of normal subjects) to yield equivalent results with the predicate device. Thirty eight normal subjects provided the samples utilized. The equivocal zone was initially determined as 16 to $24~\mu \mathrm{g / ml}$ . Analysis of 128 separate samples from normal donors indicated that the $90^{\mathrm{th}}$ percentile value was $14~\mu \mathrm{g / ml}$ . The upper $95\%$ confidence interval of the $90^{\mathrm{th}}$ percentile was $24~\mu \mathrm{g / ml}$ . The $90^{\mathrm{th}}$ percentile and the upper $95\%$ confidence limit of the $90^{\mathrm{th}}$ percentile values are similar to the chosen values of 16 to $24~\mu \mathrm{g / ml}$ for the equivocal zone.
2. Comparison studies:
a. Method comparison with predicate device:
To evaluate the performance of the assay compared with the predicate device, 81 samples were tested in both assays. The samples were identified as provided by normal asymptomatic subjects and autoimmune patients. Of the 81 samples, 38 samples were identified as normal subjects. The remaining 43 samples were identified as autoimmune patients (28 CIC, 4 SLE, 2 ANA, 1 RA, 2 TPO, 3 ENA, 1 ANCA, 1 gliadin, and 1 cardiolipin). The following table summarizes the agreement of the proposed device
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with 3 result categories (positive, equivocal, and negative) vs. the predicate device with 2 result categories (positive, negative):
SciMedx result
| IBL | >9 μg/ml | ≤ 9 μg/ml | Total |
| --- | --- | --- | --- |
| >24 μg/ml | 19 | 2 | 21 |
| 16≤ x ≤ 24 μg/ml | 1 | 0 | 1 |
| <16 μg/ml | 2 | 57 | 59 |
| total | 22 | 59 | 81 |
The amount of agreement between assay results was $93.8\%$ . The amount of chance agreement between assay results was $60.1\%$ . The kappa statistic for agreement compared with random chance agreement was $0.845 \pm 1$ standard error of 0.109. The probability of perfect agreement between assays was 0.15. Therefore, the agreement of assay results is not significantly different from perfect agreement (kappa = 1). This would support the hypothesis that the assay results are equivalent. The relative agreement of positive result was 0.864 (exact binomial $95\%$ confidence interval 0.651 to 0.971). The relative agreement of negative result was 0.966 (exact binomial $95\%$ confidence interval (0.883 to 0.996).
b. Matrix comparison:
Not performed. Only serum is used in the assay.
3. Clinical studies:
a. Clinical sensitivity:
Not performed
b. Clinical specificity:
Not performed
c. Other clinical supportive data (when $a$ and $b$ are not applicable):
Fifty-two samples from rheumatoid arthritis subjects and 45 samples from SLE subjects were purchased from a commercial supplier having an identified disease state. The mean rheumatoid factor result for the 52 subjects was $170~\mathrm{IU / ml}$ . For the 45 SLE subjects, the mean double stranded DNA (dsDNA) titer was 1:16. Among rheumatoid arthritis subjects, $51.9\%$ had assay results greater than $24~\mu \mathrm{g / ml}$ while $44.2\%$ had assay results less than $16~\mu \mathrm{g / ml}$ . Of the SLE subjects, $33.3\%$ had assay results greater than $24~\mu \mathrm{g / ml}$ while $64.4\%$ had assay results less than $16~\mu \mathrm{g / ml}$ . Samples from 129 normal blood donors were also tested in the proposed assay. Among normal blood donors, $4.7\%$ had assay results greater than $24~\mu \mathrm{g / ml}$ while $93.8\%$ had assay results less than $16~\mu \mathrm{g / ml}$ .
4. Clinical cut-off:
Not performed
5. Expected values/Reference range:
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Positive assay result > 24 µg/ml circulating immune complexes
Equivocal assay result 16 to 24 µg/ml circulating immune complexes
Negative assay result < 16 µg/ml circulating immune complexes
**M. Conclusion:**
Based on data shown in the submission, the IBL C3d–CIC Test Kit is substantially equivalent to other devices regulated under 21 CFR §866.5240, product code CZW, Class II.
