BIOTEST ANTI-EBV RECOMBINANT

K983839 · Biotest Diagnostics Corp. · LSE · Jun 9, 1999 · Microbiology

Device Facts

• Record ID: K983839
• Device Name: BIOTEST ANTI-EBV RECOMBINANT
• Applicant: Biotest Diagnostics Corp.
• Product Code: LSE · Microbiology
• Decision Date: Jun 9, 1999
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 866.3235
• Device Class: Class 1
• Attributes: Pediatric

Intended Use

The Biotest EA IgG ELISA is an enzyme immunoassay using recombinant antigens for the qualitative detection of IgG antibodies to the Epstein-Barr Virus Early Antigens (EA) p54 and p138 in human serum or plasma. Results obtained with this test, in conjunction with other clinical and patient data obtained in assays for other Epstein-Barr virus-specific antibodies such as anti-Early Antigen IgM and anti-EBNA-1 IgG, assist in serological diagnosis of EBV infection in pediatric and adult populations.

Device Story

Biotest Anti-EBV Recombinant EA IgG is an indirect ELISA test system. It uses recombinant EBV antigens (p54 and p138) immobilized on 96-well microtest plates to capture specific IgG antibodies from patient serum or plasma. After washing, captured antibodies are labeled with murine monoclonal anti-human IgG antibody-enzyme conjugates. A substrate is added, converting to a colored product; color intensity is measured via spectrophotometer at 450 nm. The test is performed in clinical laboratories. Results are interpreted by healthcare providers alongside other clinical data and EBV-specific antibody assays to aid in diagnosing EBV infection stages (e.g., acute, past, reactivation). The device provides qualitative results to assist clinical decision-making.

Clinical Evidence

Clinical study of 408 samples (pediatric and adult, ages 1-74) across various EBV infection stages. Compared against IFA and clinical interpretation. Combined site results vs. IFA: relative sensitivity 78.4%, relative specificity 97.3%. Comparison against clinical interpretation showed 98.5% sensitivity and 100% specificity. No cross-reactivity observed with HSV I/II, VZV, CMV, rheumatoid factor, ANA, or ASL positive samples. Reproducibility evaluated via inter-run, intra-run, and inter-lab testing.

Technological Characteristics

Indirect ELISA; 96-well microtest plate solid phase. Recombinant EBV antigens (p54, p138). Murine monoclonal anti-human IgG enzyme conjugate. TMB chromogen. Spectrophotometric detection at 450 nm. Manual or automated plate processing. Requires serum or plasma samples.

Indications for Use

Indicated for qualitative detection of IgG antibodies to EBV Early Antigens (p54/p138) in human serum or plasma. Used in conjunction with other clinical data and assays (e.g., EBNA IgG, EA IgM) to assist in serological diagnosis of EBV infection in pediatric and adult populations.

Regulatory Classification

Identification

Epstein-Barr virus serological reagents are devices that consist of antigens and antisera used in serological tests to identify antibodies to Epstein-Barr virus in serum. The identification aids in the diagnosis of Epstein-Barr virus infections and provides epidemiological information on diseases caused by these viruses. Epstein-Barr viruses are thought to cause infectious mononucleosis and have been associated with Burkitt's lymphoma (a tumor of the jaw in African children and young adults) and postnasal carcinoma (cancer).

Predicate Devices

• EBV-EA Test (K872617)

Submission Summary (Full Text)

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JUN 1999 1999

# ATTACHMENT I

# REVISED 510(k) SUMMARY

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# 510(k) Summary of Information Respecting Safety and Effectiveness

- A. Name and Address of Submitter

| Company Name and Address: | Biotest Diagnostics Corporation<br>66 Ford Road, Suite 131<br>Denville, NJ 07834 |
|----------------------------------------------------|----------------------------------------------------------------------------------|
| Telephone: | [phone] |
| FAX: | [phone] |
| Contact Person: | Patricia E. Bonness, Official Correspondent |
| Date 510(k) Summary was Prepared: October 26, 1998 | |
| Device Names | |
| Proprietary Name: | Biotest Anti-EBV Recombinant<br>EA IgG |
| Common Name: | EBV EA IgG |
| Classification Name: | Epstein-Barr virus serological reagents |

- C. Legally Marketed Device
B.

Biotest Diagnostics claims substantial equivalence to the EBV-EA Test (K872617) currently in commercial distribution by Gull Laboratories, Inc., Salt Lake City, UT.

#### D. Device Description

Using recombinant DNA technology, Biotest has developed three highly purified EBV antigens for use in their ELISA test system.

- EBNA-1 p72: Major antigen of the EBNA complex. The recombinant protein does not contain the glycine-alanine copolymer, a structural feature of EBNA-1, which shows cross-reactivities with certain autoantibodies and CMV (IgM response).
- EA-D p64: Early antigen. Dominant immunogen of the EA-D complex.
- EA p138: Early antigen and major DNA binding protein. This highly reactive antigen is not detectable in EA immunofluorescence assays based on chemically induced Raji cells (deletion within the Raji genome).

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| Biotest<br>Anti-EBV<br>recombinant | p72 | p54 | p134 | anti-human IgG | anti-human IgM |
|------------------------------------|-----|-----|------|----------------|----------------|
| EA IgM | | X | X | | X |
| EA IgG | | X | X | X | |
| EBNA IgG | X | | | X | |

#### Biotest Anti-EBV ELISA TESTS

The EBV immune status will be determined by the detection of specific antibodies directed against EBV proteins according to the principle of the indirect ELISA. The antigens are purified to apparent homogeneity and immobilized on the solid phase (microtest plate, 96 wells). If the patient's serum contains specific antibodies they will bind during the first incubation. Non-specific antibodies are removed by washing steps. During a second incubation the captured IgG or IgM antibodies are labeled. This is performed by addition of murine monoclonal anti-human IgG or IgM antibody-enzyme-conjugates. The final reaction converts a colorless substrate to a colored product. The concentration of color after a definite time is related to the concentration of antibody in the serum sample.

#### E. Intended Use

The Biotest EA IgG ELISA is an enzyme immunoassay using recombinant antigens for the qualitative detection of IgG antibodies to the Epstein-Barr Virus Early Antigens (EA) p54 and p138 in human serum or plasma. Results obtained with this test, in conjunction with other clinical and patient data obtained in assays for other Epstein-Barr virus-specific antibodies such as anti-Early Antigen IgM and anti-EBNA-1 IgG, assist in serological diagnosis of EBV infection in pediatric and adult populations.

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- F. Comparison with Predicate Device
A summary comparison of the features of the Biotest EA IgG and the Gull EA IgG test kits is provided in Table 1 below:

# Table 1

Feature Comparison of Biotest and Gull EBV-EA Test Kits

| | Biotest | Gull |
|------------------------------------------|---------------------------------------------------------------------------------------------|----------------------------------------------------------------------------|
| Intended Use | Detection of IgG antibodies<br>to EBV EA<br>Qualitative only | Detection of IgG antibodies<br>to EBV EA<br>Qualitative and quantitative |
| Assay Method | ELISA | Indirect fluorescent antibody (IFA) |
| Reactive Ingredients | Recombinant EBV (EA p54/p138)<br>Peroxidase-conjugated monoclonal<br>anti-human IgG (mouse) | EBV infected Raji cells<br>Fluorescein-labeled<br>anti-human IgG (caprine) |
| Specimen: Type<br>Min. Volume<br>Storage | Serum or plasma<br>25 µl<br>2 - 8°C or -20°C | Serum<br>15 µl of 1:10 dilution<br>2 - 8°C/7 days or -20°C |
| Controls | Negative<br>Positive | Negative<br>Positive |
| Chromogen | TMB | Fluorescein; Evans blue |
| Results:<br>Evaluation | spectrophotometer @ 450 nm | Fluorescent microscope |
| Kit Size | 96 tests | 100 tests |

୧ ୦

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#### G. Performance Data

#### Sensitivity/Specificity

The performance of the Biotest EA IgG ELISA was evaluated in a clinical study of 408 patient samples conducted at two geographically distinct locations. Samples were obtained from both pediatric and adult patients (ages 1 to 74) representing acute (139), late acute (34), recent past (12), past (120), reactivation (22), past/probable reactivation (11), and negative (70) disease stages of EBV infection.

Two methods were used to evaluate the performance of the Biotest EA IgG ELISA: direct comparison with commercially available or published EA IgG IFA tests, and comparison with clinical interpretation (stage of infection based on antibody patterns and clinical diagnosis at the time the specimen was drawn).

Results of the direct comparison with IFA for both sites combined demonstrated a relative sensitivity of 78.4% and a relative specificity of 97.3%.

#### Table 1

### Clinical Site 1 Direct Comparison to EA IgG IFA

| EA IgG IFA | | | |
|------------|-----|----|-------|
| | + | - | Total |
| Biotest + | 125 | 0 | 125 |
| Biotest - | 32 | 51 | 83 |
| Total | 157 | 51 | 208 |

Relative Sensitivity = 79.6% Relative Specificity = 100% Relative Agreement = 84.6% (C.I. = 73.3 to 85.9%) (C.I. = 93.0 to 100%)

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#### Table 2

| EA IgG IFA | | | |
|------------|-----|----|-------|
| | + | - | Total |
| Biotest + | 107 | 3 | 110 |
| Biotest - | 32 | 58 | 90 |
| Total | 139 | 61 | 200 |

# Clinical Site 2 Direct Comparison to EA IgG IFA

・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・ Relative Sensitivity = 77.0% Relative Specificity = 95.1% Relative Agreement = 82.5%

(C.I. = 70 to 84%) (C.I. = 86.3 to 99%)

### Table 3

### Combined Site Results Direct Comparison to EA IgG

| EA IgG IFA | | | |
|------------|-----|-----|-------|
| | + | - | Total |
| Biotest + | 232 | 3 | 235 |
| Biotest - | 64 | 109 | 173 |
| Total | 296 | 112 | 408 |

Relative Sensitivity = 78.4% Relative Specificity = 97.3% Relative Agreement = 83.6%

------------------------------------------------------------------------------------------------------------------------------------------------------------------------------

(C.I. = 73.7 to 83.1%) (C.I. = 92.4 to 99.4%)

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Results based on Clinical Interpretation of all patient samples where Biotest ELISA EA IgG antibody responses matched expected serological pattern analysis for each state of infection, including - Acute, Convalescent (Late Acute or Recent Past), Past, Reactivation and Probable Reactivation/past, and Negative.

| Clinical Interpretation | | | | | |
|--------------------------------------------|----------------------------|-----------------------------|----------------------------|-----------------------------|---------------------------|
| | Acute | Convalescent | Past | Reactivation | Negative |
| Result matches<br>Biotest | 138 | 44 | 119 | 32 | 0 |
| Result does not<br>match Biotest | 1 | 2 | 1 | 1 | 70 |
| Sensitivity<br>Specificity<br>95% to C.I.% | 99.2%<br>N/A<br>96.1 - 100 | 95.6%<br>N/A<br>85.2 - 99.5 | 99.2%<br>N/A<br>95.4 - 100 | 97.0%<br>N/A<br>84.2 - 99.9 | N/A<br>100%<br>94.9 - 100 |

Total Clinical Sensitivity = 98.5% Total Clinical Specificity = 100% Total Clinical Agreement = 98.7% (C.I. = 96.6 to 99.5%) (C.I. = 94.9 to 100%)

Note: C.I. = 95% confidence intervals calculated by the exact method.

### Cross Reactivity

ﻟﺴﻴﻨﺎ

No cross reactivity was observed when the Biotest EA IgG ELISA was used to test samples from patients acutely infected with:

| Herpes Simplex Virus I/II | n = 33 |
|---------------------------|--------|
| Varicella Zoster Virus | n = 42 |
| Cytomegalovirus | n = 12 |

Further, no cross reactivity was observed in tests of rheumatoid factor positive (n = 10), antinuclear antibody positive (n = 5), and ASL positive (n = 15) samples

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## Reproducibility

ﻟ

To evaluate the reproducibility of the Biotest EA IgG ELISA, a panel of 10 patient serum specimens (low to high positive) was tested at the clinical sites. The mean, standard deviation (S.D.) and coefficient of variation (C.V.) for inter-run, intra-run and inter-lab reproducibility are presented below.

| | Inter-Run (n = 10) | | | Intra-Run (n = 8) | | |
|---------|--------------------|-------|--------|-------------------|-------|--------|
| Panel # | Mean | S.D. | % C.V. | Mean | S.D. | % C.V. |
| 1 | 2.186 | 0.167 | 7.6 | 2.209 | 0.123 | 5.6 |
| 2 | 1.871 | 0.143 | 7.6 | 2.083 | 0.108 | 5.2 |
| 3 - | 2.169 | 0.311 | 14.3 | 2.332 | 0.157 | 6.7 |
| 4 | 0.744 | 0.077 | 10.3 | 0.600 | 0.057 | 9.5 |
| 5 | 2.208 | 0.366 | 16.6 | 2.317 | 0.306 | 13.2 |
| 6 | 1.503 | 0.193 | 12.9 | 1.343 | 0.057 | 4.3 |
| 7 | 2.488 | 0.308 | 12.4 | 2.185 | 0.116 | 5.3 |
| 8 | 2.288 | 0.218 | 9.5 | 2.685 | 0.272 | 10.1 |
| 9 | 2.386 | 0.341 | 14.3 | 2.712 | 0.192 | 7.1 |
| 10 | 2.201 | 0.269 | 12.2 | 2.299 | 0.090 | 3.9 |

Site #1

Site #2

| | Inter-Run (n = 4) | | | Intra-Run (n = 24) | | |
|---------|-------------------|-------|--------|--------------------|-------|--------|
| Panel # | Mean | S.D. | % C.V. | Mean | S.D. | % C.V. |
| 1 | 1.783 | 0.311 | 17.4 | 2.134 | 0.409 | 19.2 |
| 2 | 1.235 | 0.177 | 14.3 | 1.413 | 0.269 | 19.0 |
| 3 | 2.168 | 0.265 | 12.2 | 2.504 | 0.381 | 15.2 |
| 4 | 0.619 | 0.060 | 9.8 | 0.832 | 0.130 | 15.7 |
| 5 | 2.003 | 0.183 | 9.1 | 2.530 | 0.408 | 16.1 |
| 6 | 1.215 | 0.121 | 10.0 | 1.733 | 0.244 | 14.1 |
| 7 | 1.929 | 0.163 | 8.4 | 2.580 | 0.425 | 16.5 |
| 8 | 1.742 | 0.167 | 9.6 | 2.412 | 0.408 | 16.9 |
| 9 | 2.604 | 0.209 | 8.0 | 2.285 | 0.379 | 16.6 |
| 10 | 1.503 | 0.152 | 10.1 | 2.066 | 0.247 | 12.0 |

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Inter-Lab (n = 14)

ﻓﻲ ﺍﻟﻤﺴﺘﻘﻠﺔ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍ

| Panel # | Mean | S.D. | % C.V. |
|---------|-------|-------|--------|
| 1 | 2.076 | 0.277 | 13.3 |
| 2 | 1.696 | 0.335 | 19.7 |
| 3 | 2.176 | 0.276 | 12.7 |
| 4 | 0.719 | 0.101 | 14.1 |
| 5 | 2.136 | 0.316 | 14.8 |
| 6 | 1.395 | 0.221 | 15.9 |
| 7 | 2.302 | 0.366 | 15.9 |
| 8 | 2.125 | 0.315 | 14.8 |
| 9 | 2.444 | 0.305 | 12.5 |
| 10 | 2.003 | 0.397 | 19.8 |

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Image /page/9/Picture/1 description: The image shows the logo for the Department of Health & Human Services - USA. The logo consists of a circular border with the text "DEPARTMENT OF HEALTH & HUMAN SERVICES - USA" written around it. Inside the circle is an abstract image of an eagle.

Public Health Service

JUN 9 1999 Food and Drug Administration 9200 Corporate Boulevard Rockville MD 20850

Ms. Patricia E. Bonness Official Correspondent Biotest Diagnostics Corporation 66 Ford Road Suite 131 Denville. New Jersey 07834

Re: K983839

Trade Name: Biotest Anti-EBV Recombinant EA IgG Regulatory Class: I Product Code: LSE Dated: March 15, 1999 Received: March 31, 1999

Dear Ms. Bonness:

We have reviewed your Section 510(k) notification of intent to market the device referenced above and we 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). 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.

If your device is classified (see above) into either class II (Special Controls) or class III (Premarket Approval), it may be subject to such additional controls. Existing major regulations affecting your device can be found in the Code of Federal Regulations, Title 21, Parts 800 to 895. A substantially equivalent determination assumes compliance with the Current Good Manufacturing Practice requirements, as set forth in the Quality System Regulation (QS) for Medical Devices: General regulation (21 CFR Part 820) and that, through periodic QS inspections, the Food and Drug Administration (FDA) will verify such assumptions. Failure to comply with the GMP regulation may result in regulatory action. In addition, FDA may publish further announcements concerning your device in the Federal Register. Please note: this response to your premarket notification submission does not affect any obligation you might have under sections 531 through 542 of the Act for devices under the Electronic Product Radiation Control provisions, or other Federal laws or regulations.

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#### Page 2

Under the Clinical Laboratory Improvement Amendments of 1988 (CLIA-88), this device may require a CLIA complexity categorization. To determine if it does, you should contact the Centers for Disease Control and Prevention (CDC) at [phone].

This letter will allow you to begin marketing your device as described in your 510(k) premarket notification. The FDA finding of substantial equivalence of your device to a legally marketed predicate device results in a classification for your device and thus, permits your device to proceed to the market.

If you desire specific advice for your device on our labeling regulation (21 CFR Part 801 and additionally 809.10 for in vitro diagnostic devices), please contact the Office of Compliance at [phone]. Additionally, for questions on the promotion and advertising of your device, please contact the Office of Compliance at [phone]. Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR 807.97). Other general information on your responsibilities under the Act may be obtained from the Division of Small Manufacturers Assistance at its toll free number [phone] or at [phone] or at its internet address "http://www.fda.gov/cdrh/dsmamain.html"

Sincerely yours,

Steven Sutman

Steven I. Gutman, M.D., M.B.A. Director Division of Clinical Laboratory Devices Office of Device Evaluation Center for Devices and Radiological Health

Enclosure

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Page ***_*** of **_**

510(k) Number (if known):

Device Name:__________________________________________________________________________________________________________________________________________________________________ Biotest Anti-EBV Recombinant EA IgG

Indications For Use:

The Biotest EA IgG is an enzyme immunoassay for the detection of IgG antibodies to the Epstein-Barr Virus (EBV-EA (Early Antigens) p54/p138) in human serum or plasma.

It is indicated for use, in conjunction with other clinical and patient data obtained in assays for other Epstein-Barr antigens such as EBNA IgG and Early Antigen IgM, in the serological diagnosis of EBV infection.

# (PLEASE DO NOT WRITE BELOW THIS LINE-CONTINUE ON ANOTHER PAGE IF NEEDED)

Concurrence of CDRH, Office of Device Evaluation (ODE)

Prescription Use (Per 21 CFR 801.109)

OR

Over-The-Counter Use

(Optional Format 1-2-96)