← Product Code [NHX](/submissions/IM/subpart-f%E2%80%94immunological-test-systems/NHX) · K072944

# QUANTA LITE CCP3.1 IGG/IGA ELISA (K072944)

_Inova Diagnostics, Inc. · NHX · Mar 5, 2008 · Immunology · SESE_

**Canonical URL:** https://fda.innolitics.com/submissions/IM/subpart-f%E2%80%94immunological-test-systems/NHX/K072944

## Device Facts

- **Applicant:** Inova Diagnostics, Inc.
- **Product Code:** [NHX](/submissions/IM/subpart-f%E2%80%94immunological-test-systems/NHX.md)
- **Decision Date:** Mar 5, 2008
- **Decision:** SESE
- **Submission Type:** Traditional
- **Regulation:** 21 CFR 866.5775
- **Device Class:** Class 2
- **Review Panel:** Immunology

## Intended Use

The QUANTA Lite™ CCP3.1 IgG/IgA ELISA is a semiquantitative enzyme-linked immunosorbent assay for the detection of IgG and IgA anti-CCP3 (Cyclic Citrullinated Peptide 3) antibodies in patient sera or citrated or EDTA plasma. The presence of these antibodies, when considered in conjunction with other laboratory and clinical findings, is an aid in the diagnosis of rheumatoid arthritis (RA), including RA diagnosed within 2 years of presentation of symptoms.

## Device Story

The QUANTA Lite™ CCP3.1 IgG/IgA ELISA is an in vitro diagnostic test used in clinical laboratories. It utilizes patient serum or plasma samples to detect IgG and IgA antibodies against Cyclic Citrullinated Peptide 3 (CCP3). The assay follows an enzyme-linked immunosorbent assay (ELISA) format. Results are interpreted by healthcare professionals alongside other clinical and laboratory findings to assist in diagnosing rheumatoid arthritis. The device provides a semiquantitative measurement, aiding in the identification of patients with RA, including those in early stages of the disease.

## Clinical Evidence

No clinical data provided in the document.

## Technological Characteristics

Semiquantitative enzyme-linked immunosorbent assay (ELISA) for detection of IgG and IgA anti-CCP3 antibodies. Analyte: Cyclic Citrullinated Peptide 3. Sample types: serum, citrated plasma, or EDTA plasma.

## Regulatory Identification

A rheumatoid factor immunological test system is a device that consists of the reagents used to measure by immunochemical techniques the rheumatoid factor (antibodies to immunoglobulins) in serum, other body fluids, and tissues. Measurement of rheumatoid factor may aid in the diagnosis of rheumatoid arthritis.

## Submission Summary (Full Text)

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>
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# 510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION DECISION SUMMARY

A. 510(k) Number:
k072944

B. Purpose for Submission:
New device

C. Measurand:
Anti-cyclic citrullinated peptide 3 (CCP3)

D. Type of Test:
Semiquantitative enzyme-linked immunosorbent assay (ELISA)

E. Applicant:
INOVA Diagnostics, Inc.

F. Proprietary and Established Names:
QUANTA Lite™ CCP3.1 IgG/IgA ELISA

G. Regulatory Information:

|  Product Code | Classification | Regulation Section | Panel  |
| --- | --- | --- | --- |
|  NHX – Antibodies, anti-cyclic citrullinated peptide | Class II | 21 CFR 866.5775
Rheumatoid factor immunological test system | Immunology 82  |

H. Intended Use:

1. Intended use(s):
The QUANTA Lite CCP3.1 IgG/IgA ELISA is a semiquantitative enzyme-linked immunosorbent assay for the detection of IgG and IgA anti-CCP3 (Cyclic Citrullinated Peptide 3) antibodies in patient sera or citrated or EDTA plasma. The presence of these antibodies, when considered in conjunction with other laboratory and clinical findings, is an aid in the diagnosis of rheumatoid arthritis (RA), including RA diagnosed within 2 years of presentation of symptoms.

2. Indication(s) for use:
Same as Intended Use

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

4. Special instrument requirements:
Microplate reader capable of measuring OD at 450/620 nm

I. Device Description:
The QUANTA Lite CCP3.1 IgG/IgA ELISA consists of a polystyrene microwell ELISA plate coated with purified, synthetic CCP3 antigen; ELISA negative, low positive and high positive (Calibrator A) controls; calibrators A–E (250, 125, 62.5, 31.25 and 15.62 Units); sample diluent; high specificity wash concentrate; goat anti-human IgG/IgA horseradish peroxidase conjugate; TMB chromogen; and stop solution.

J. Substantial Equivalence Information:

1. Predicate device name(s):

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QUANTA Lite™ CCP IgG ELISA

2. Predicate 510(k) number(s):
k020414

3. Comparison with predicate:

|  Similarities  |   |   |
| --- | --- | --- |
|  Item | Device | Predicate  |
|   | CCP3.1 IgG/IgA ELISA | CCP IgG ELISA  |
|  Indications for Use | To aid in the diagnosis of rheumatoid arthritis, including RA diagnosed within 2 years of presentation of symptoms | To aid in the diagnosis of rheumatoid arthritis  |
|  Method | ELISA | Same  |
|  Serum dilution and volume | 1:101; 100 μl | Same  |
|  Calculation of results | Ratio method: compared to low control unit value; Standard curve: unknown concentrations read from plot of calibrators | Same  |
|  Result interpretation | <20 U/mL = negative, 20 - 39 = weak positive, 40 - 59 = mod. positive ≥ 60 = strong positive | Same  |
|  Units | Arbitrary ELISA units | Same  |
|  Solid phase | Coated polystyrene microwell plates | Same  |
|  Sample diluent | Tris-buffered saline, Tween 20, absorbents and protein stabilizers | Same  |
|  Controls | Negative, low positive, and high positive | Same  |
|  Differences  |   |   |
| --- | --- | --- |
|  Item | Device | Predicate  |
|   | CCP3.1 IgG/IgA ELISA | CCP IgG ELISA  |
|  Analyte measured | Anti-cyclic citrullinated peptide 3 | Anti-cyclic citrullinated peptide 2  |
|  Capture antigen | Synthetic CCP3 | Synthetic CCP2  |
|  Conjugate | HRP conjugated goat anti-human IgG/IgA | HRP conjugated goat anti-human IgG  |
|  Test matrix | Serum and citrated or EDTA plasma | Serum  |
|  Wash concentrate | 10X High specificity | 40X Tris-buffered saline and Tween 20  |

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K. Standard/Guidance Document Referenced (if applicable):
None referenced

L. Test Principle:
The assay utilizes plastic microwells as a solid phase for attachment of the synthetic cyclic citrullinated peptide. Pre-diluted controls and diluted patient samples are added to separate wells, allowing any CCP IgG and/or IgA antibodies present to bind to the immobilized antigen. Unbound sample is washed away and an enzyme labeled anti-human IgG/IgA conjugate is added to each well. A second incubation allows the enzyme labeled anti-human IgG/IgA to bind to any patient antibodies that have become attached to the microwells. After washing away any unbound enzyme labeled anti-human IgG/IgA, the remaining enzyme activity is measured by adding a chromogenic substrate and measuring the intensity of the color that develops. The assay is evaluated by spectrophotometrically measuring and comparing the color intensity that develops in the patient wells with the color in the control wells (ratio). Alternatively, calibrator absorbance results are plotted and unknowns interpreted by reading the corresponding absorbance from the standard curve and calculating the units. Results determined with the assay are interpreted as negative, weak positive, moderately positive, or strong positive and are reported in arbitrary units.

M. Performance Characteristics (if/when applicable):

1. Analytical performance:
a. Precision/Reproducibility:
Intra-assay performance for the assay was measured by running 9 samples 9 or 10 times each on the same ELISA plate. Percent CVs ranged from 3-13%. Representative results are shown below.

|   | Negative |   | Low 1 |   | Low 2 |   | High 1  |   |
| --- | --- | --- | --- | --- | --- | --- | --- | --- |
|   |  1 pt.* | 5 pt.** | 1 pt. | 5 pt. | 1 pt. | 5 pt. | 1 pt. | 5 pt.  |
|  Mean units | 4 | 2 | 22 | 24 | 26 | 30 | 68 | 81  |
|  SD | 0.2 | 0.2 | 0.9 | 1.1 | 0.9 | 1.0 | 1.9 | 3.2  |
|  CV% | 4% | 13% | 4% | 5% | 3% | 3% | 3% | 4%  |

* Ratio method test results; ** Standard curve test results

Inter-assay performance for the assay was determined by testing 16 samples: negative, low positive and strong positive samples in 6 separate assays spanning up to 9 different days. Overall percent CVs ranged from 1 to 11%. Representative results are shown below.

|   | Negative |   | Low 1 |   | Low 2 |   | High 1  |   |
| --- | --- | --- | --- | --- | --- | --- | --- | --- |
|   | 1 pt.* | 5 pt.** | 1 pt. | 5 pt. | 1 pt. | 5 pt. | 1 pt. | 5 pt.  |
|  Mean units | 6 | 4 | 22 | 23 | 28 | 30 | 69 | 85  |
|  SD | 0.4 | 0.5 | 0.7 | 0.6 | 0.8 | 1.2 | 1.7 | 1.2  |
|  CV% | 8% | 11% | 3% | 3% | 3% | 4% | 2% | 1%  |

* Ratio method test results; ** Standard curve test results

b. Linearity/assay reportable range:

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No claims were made regarding linearity for the assay. Reactivity is related to the quantity of antibody present in a non-linear fashion. While increases and decreases in patient antibody concentration will be reflected in a corresponding rise or fall in reactivity, the change is not proportional (i.e. a doubling of the antibody concentration will not double the reactivity). Specimens giving OD readings above the readable range of the plate reader may be reported as greater than the highest measurable OD. The assay has the option of interpretation of results from a standard curve but results are still semi-quantitative. Values calculated with the ratio method will be different than those calculated by the standard curve. High values will differ more than low values. The CCP3.1 IgG/IgA High Positive will not yield a value of 250 Units with the ratio method.

c. Traceability, Stability, Expected values (controls, calibrators, or methods): There is no recognized standard reference material for CCP. Based on results from accelerated stability testing, components were assigned expiration date of one year.

d. Detection limit: The determination of detection limit/analytical sensitivity was not performed or relevant for this assay.

e. Analytical specificity: Interfering substances: To assess the possibility of false positive results from high levels of hemoglobin, bilirubin, cholesterol or triglycerides, sera with known quantities of the potential interferant were mixed with normal sera and run on the assay. To evaluate the potential for false negative results, the substances were mixed with a positive CCP3.1 IgG/IgA serum.

|  Substance added | Normal serum (Units) | Result Interpretation | Positive serum (Units) | % Difference  |
| --- | --- | --- | --- | --- |
|  None | 5 | Negative | 62 |   |
|  Hemoglobin 1000 mg/dL | 10 | Negative | 59 | 4.8  |
|  Bilirubin 29.7 mg/dL | 7 | Negative | 56 | 9.7  |
|  Cholesterol 354 mg/dL | 5 | Negative | 54 | 12.9  |
|  Triglycerides 1016 mg/dL | 6 | Negative | 58 | 6.5  |

For samples closer to the assay cut-off (25-40 Units), % recoveries ranged from 84-110% for the single point and 87-112% for the 5 point curve interpretations.

Cross-reactivity To assess potential cross-reactivity with other autoantibodies, the new assay was evaluated with 16 samples containing high levels of various other autoantibodies. Included in this group were 2 samples each that were positive

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for SS-A, SS-B, Sm, RNP, Scl-70, Jo-1, ribosomal P, and dsDNA autoantibodies. All samples were negative for anti-CCP3.1 antibodies.

f. Assay cut-off:

To determine the reference interval (cut-off) a total of 275 samples from random blood donors were tested. At a cut-off of  $\geq 20$  Units, 2 samples were positive for a specificity of  $99.3\%$ .

# 2. Comparison studies:

a. Method comparison with predicate device:

Testing was performed on 942 specimens that included 275 from healthy individuals; 495 rheumatoid arthritis (RA) patient sera including 86 RA (presented with symptoms  $&lt; 2$  years); 74 other rheumatic disease patient sera (24 scleroderma, 16 Sjögren's syndrome, and 34 SLE samples); and 98 infectious disease sera specimens (58 HCV, 14 HSV, 14 CMV6 toxoplasmosis and 6 rubella).

|   | INOVA QUANTA Lite IgG ELISA  |   |   |   |
| --- | --- | --- | --- | --- |
|   |   |  Positive | Negative | Total  |
|  INOVA QUANTA | Positive | 320 | 38* | 358  |
|  Lite CCP3.1 | Negative | 8** | 576 | 584  |
|  IgG/IgA ELISA | Total | 328 | 614 | 942  |

* 33 of these samples are diagnosed RA
** 5 of these samples are diagnosed RA

Positive percent agreement:  $320 / 328 = 97.6\%$  95% CI: 95.3-98.9%

Negative percent agreement:  $576 / 614 = 93.8\%$  95% CI: 91.6-95.6%

Overall agreement:  $896 / 942 = 95.1\%$

b. Matrix comparison:

Seven serum, EDTA plasma, and citrated plasma were serially diluted to demonstrate that the matrices showed similar results. The comparisons are shown below. Serum versus EDTA plasma yielded  $y = 0.9971x + 12.012$ ,  $R^2 = 0.9877$ .

![img-0.jpeg](img-0.jpeg)

Serum versus citrated plasma yielded  $y = 0.9494x - 0.3502$ ,  $R^2 = 0.9945$ .

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![img-1.jpeg](img-1.jpeg)

3. Clinical studies:

a. Clinical Sensitivity and Specificity:

The clinical studies involved testing 942 specimens that included 275 from healthy individuals; 495 rheumatoid arthritis (RA) patient sera including 86 early RA (presented with symptoms &lt;2 years); 74 other rheumatic disease patient sera; and 98 infectious disease sera specimens. The following results were obtained:

|   | Rheumatoid Arthritis  |   |   |   |
| --- | --- | --- | --- | --- |
|   |   |  Positive | Negative | Total  |
|  INOVA QUANTA | Positive | 348 | 10 | 358  |
|  Lite CCP3.1 | Negative | 147 | 437 | 584  |
|  IgG/IgA ELISA | Total | 495 | 447 | 942  |

Clinical sensitivity: 348/495 = 70.3% 95% CI: 66.1-74.3%

Clinical specificity: 437/447 = 97.8% 95% CI: 95.9-98.9%

Overall agreement: 785/942 = 83.3%

The clinical sensitivity for the 86 early RA patients was 55/86 (64%).

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

Not applicable

4. Clinical cut-off:

See assay cut-off

5. Expected values/Reference range:

The expected value in the normal population is negative. However, the presence of autoantibodies related to RA increases with age. Using the established cut-off of &lt;20 Units as negative, in a study group of 272 normal blood donors, 99.3% were negative for the presence of anti-CCP3.1 autoantibodies. There are suggested cut-offs in the labeling with a recommendation that each laboratory establish its own normal range.

N. Proposed Labeling:

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

**O. Conclusion:**

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

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**Source:** [https://fda.innolitics.com/submissions/IM/subpart-f%E2%80%94immunological-test-systems/NHX/K072944](https://fda.innolitics.com/submissions/IM/subpart-f%E2%80%94immunological-test-systems/NHX/K072944)

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