BIOTEST ANTI-EBV RECOMBINANT, EA IGM

K983842 · Biotest Diagnostics Corp. · LSE · Jun 9, 1999 · Microbiology

Device Facts

• Record ID: K983842
• Device Name: BIOTEST ANTI-EBV RECOMBINANT, EA IGM
• 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 IgM ELISA is an enzyme immunoassay using recombinant antigens for the qualitative detection of IgM 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 IgG and anti-EBNA-1 IgG, assist in serological diagnosis of EBV infection in pediatric and adult populations.

Device Story

Biotest EA IgM ELISA is an in vitro diagnostic test for qualitative detection of IgM antibodies to Epstein-Barr Virus (EBV) early antigens (p54 and p138) in human serum or plasma. Device uses recombinant DNA technology to produce highly purified antigens immobilized on 96-well microtest plates. Principle of operation is indirect ELISA: patient serum is incubated with immobilized antigens; specific IgM antibodies bind; non-specific antibodies are washed away; murine monoclonal anti-human IgM antibody-enzyme-conjugates are added; final reaction with substrate produces color. Color intensity, measured via spectrophotometer at 450 nm, is proportional to antibody concentration. Used in clinical laboratories by trained personnel. Results are interpreted by clinicians alongside other EBV-specific antibody assays (e.g., anti-EA IgG, anti-EBNA-1 IgG) and patient clinical data to assist in diagnosing EBV infection stages (acute, past, reactivation).

Clinical Evidence

Clinical study of 408 patient samples (pediatric and adult, ages 1-74) across two sites. Evaluated against VCA IgM/IgG IFA and clinical interpretation of infection stage. Combined direct comparison to IFA showed 84.1% relative sensitivity and 89.4% relative specificity. Clinical interpretation comparison showed 98.2% sensitivity and 97.1% specificity. Cross-reactivity testing performed on samples positive for HSV I/II, VZV, CMV, Toxoplasma, rheumatoid factor, ANA, and ASL; no cross-reactivity observed. Reproducibility evaluated via inter-run, intra-run, and inter-lab testing with 10 patient serum specimens.

Technological Characteristics

ELISA-based immunoassay; utilizes recombinant EBV antigens (p54, p138) immobilized on 96-well microtest plates; detection via murine monoclonal anti-human IgM antibody-enzyme-conjugate (HRP); chromogen TMB; spectrophotometric readout at 450 nm; requires serum or plasma samples; storage at 2-8°C or -20°C.

Indications for Use

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

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 IgM Test (K822734)

Submission Summary (Full Text)

{0}------------------------------------------------

JUN g 1999

:

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

K983842

# ATTACHMENT I

# REVISED 510(k) SUMMARY

57

{1}------------------------------------------------

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

- A. Name and Address of Submitter Biotest Diagnostics Corporation Company Name and Address: 66 Ford Road, Suite 131 Denville, NJ 07834 [phone] Telephone: [phone] FAX: Patricia E. Bonness, Official Correspondent Contact Person: Date 510fk) Summary was Prepared: October 26, 1998 B. Device Names Biotest Anti-EBV Recombinant Proprietary Name: EA IgM EBV EA IgM Common Name: Epstein-Barr virus serological reagents Classification Name:
- C. Legally Marketed Device

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

-

Biotest Diagnostics claims substantial equivalence to the EBV IgM Test (K822734) 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).

{2}------------------------------------------------

| Biotest<br>Anti-EBV<br>recombinant | recombinant antigens | | | monoclonal secondary antibody (HRP-conjugated) | |
|------------------------------------|----------------------|-----|------|------------------------------------------------|----------------|
| | 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 IgM antibodies are labeled. This is performed by addition of murine monoclonal anti-human 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 IgM ELISA is an enzyme immunoassay using recombinant antigens for the qualitative detection of IgM 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 IgG and anti-EBNA-1 IgG, assist in serological diagnosis of EBV infection in pediatric and adult populations.

{3}------------------------------------------------

- Comparison with Predicate Device F.
:

।

(

ﻧﮯ

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

A summary comparison of the features of the Biotest EA IgM and the Gull VCA IGM test kits is provided in Table 1 below:

## Table 1 Feature Comparison of Biotest and Gull EBV IgM Test Kits

| | Biotest | Gull |
|------------------------------------------|---------------------------------------------------------------------------------------------|---------------------------------------------------------------------------|
| Intended Use | Detection of IgM antibodies<br>to EBV EA<br>Qualitative only | Detection of IgM antibodies<br>to EBV VCA<br>Qualitative only |
| Assay Method | ELISA | Indirect fluorescent antibody (IFA) |
| Reactive Ingredients | Recombinant EBV (EA p54/p138)<br>Peroxidase-conjugated monoclonal<br>anti-human IgM (mouse) | HR1, Burkitt's lymphocytic cells<br>Fluorescein-labeled<br>anti-human IgM |
| 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 and 1:40 dilutions<br>2 - 8°C/7days or -20°C |
| Controls | Negative<br>Positive | Negative<br>Positive |
| Chromogen | TMB | Fluorescein |
| Results:<br>Evaluation | spectrophotometer @ 450 nm | Fluorescence microscope |
| Kit Size | 96 tests | 50 or 100 tests |

୧୦

{4}------------------------------------------------

G. Performance Data

#### Sensitivity/Specificity

-

The performance of the Biotest EA IgM 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 IgM ELISA: direct comparison with commercially available or published EA IgM 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 VCA IgM/IgG IFA for both sites combined demonstrated a relative sensitivity of 84.1% and a relative specificity of 89.4%.

#### Table 1

### Clinical Site 1 Direct Comparison to VCA IgM/IgG IFA

| VCA IgM/IgG IFA | | | |
|-----------------|----|-----|-------|
| | + | - | Total |
| Biotest + | 73 | 13 | 86 |
| Biotest - | 14 | 108 | 122 |
| Total | 87 | 121 | 208 |

Relative Sensitivity = 83.9% Relative Specificity = 89.3% Relative Agreement = 87.0% (C.I. = 74.5 to 90.9%) (C.I. = 83.7 to 94.8%)

{5}------------------------------------------------

### Table 2

| VCA IgM/IgG IFA | | | |
|-----------------|----|-----|-------|
| | + | - | Total |
| Biotest + | 80 | 11 | 91 |
| Biotest - | 15 | 94 | 109 |
| Total | 95 | 105 | 200 |

## Clinical Site 2 Direct Comparison to VCA IgM/IgG IFA

《《 Relative Sensitivity = 84.2% Relative Specificity = 89.5% Relative Agreement = 87.0%

..............................................................................................................................................................................

)

1

(C.I. = 75.3 to 90.0%) (C.I. = 83.7 to 95.4%)

## Table 3

## Combined Site Results Direct Comparison to VCA IgM/IgG

| VCA IgM/IgG | | | |
|-------------|-----|-----|-------|
| | + | - | Total |
| Biotest + | 153 | 24 | 177 |
| Biotest - | 29 | 202 | 231 |
| Total | 182 | 226 | 408 |

Relative Sensitivity = 84.1% Relative Specificity = 89.4% Relative Agreement = 87.0%

(C.I. = 78.7 to 89.4%) (C.I. = 85.4 to 93.4%)

{6}------------------------------------------------

Results based on Clinical Interpretation of all patient samples where Biotest ELISA EA IgM 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 | 45 | 117 | 32 | 2 |
| Result does not<br>match Biotest | 1 | 1 | 3 | 1 | 68 |
| Sensitivity<br>Specificity<br>95% to C.I.% | 99.2%<br>N/A<br>96.1 - 100 | 97.8%<br>N/A<br>88.5 - 99.9 | 97.5%<br>N/A<br>92.9 - 99.5 | 96.9%<br>N/A<br>84.2 - 99.9 | N/A<br>97.1%<br>90.1 - 99.7 |

Total Clinical Sensitivity = 98.2% Total Clinical Specificity = 97.1% Total Clinical Agreement = 98.0% (C.I. = 96.2 to 99.3%) (C.I. = 90.1 to 99.7%)

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

### Cross Reactivity

-

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

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

| Herpes Simplex Virus I/II | 4 of 5 samples found non-reactive |
|---------------------------|------------------------------------|
| Varicella Zoster Virus | 5 of 11 samples found non-reactive |
| Cytomegalovirus | 3 of 9 samples found non-reactive |
| Toxoplasma | 4 of 5 samples found non-reactive |

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.

{7}------------------------------------------------

## Reproducibility

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

ﻜﺴﺎ

To evaluate the reproducibility of the Biotest EA IgM 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 = 11) | | | Intra-Run (n = 8) | | |
|---------|--------------------|-------|--------|-------------------|-------|--------|
| Panel # | Mean | S.D. | % C.V. | Mean | S.D. | % C.V. |
| 1 | 1.799 | 0.175 | 9.7 | 1.801 | 0.065 | 3.6 |
| 2 | 1.013 | 0.171 | 16.9 | 1.140 | 0.032 | 2.8 |
| 3 - | 1.280 | 0.183 | 14.3 | 1.286 | 0.070 | 5.5 |
| 4 | 0.294 | 0.033 | 11.2 | 0.287 | 0.018 | 6.3 |
| 5 | 1.065 | 0.119 | 11.2 | 1.189 | 0.059 | 5.0 |
| 6 | 0.651 | 0.088 | 13.5 | 0.680 | 0.034 | 5.1 |
| 7 | 1.231 | 0.152 | 12.3 | 1.325 | 0.060 | 4.5 |
| 8 | 0.959 | 0.130 | 13.5 | 0.946 | 0.060 | 6.3 |
| 9 | 0.798 | 0.100 | 12.5 | 0.811 | 0.037 | 4.6 |
| 10 | 0.664 | 0.067 | 10.0 | 0.711 | 0.020 | 2.9 |

Site #1

Site #2

| | Inter-Run (n = 5) | | | Intra-Run (n = 24) | | |
|---------|-------------------|-------|--------|--------------------|-------|--------|
| Panel # | Mean | S.D. | % C.V. | Mean | S.D. | % C.V. |
| 1 | 1.641 | 0.127 | 7.7 | 1.829 | 0.268 | 14.7 |
| 2 | 1.165 | 0.036 | 3.1 | 1.257 | 0.164 | 13.1 |
| 3 | 1.415 | 0.043 | 3.1 | 1.471 | 0.124 | 8.4 |
| 4 | 0.335 | 0.020 | 6.0 | 0.362 | 0.052 | 14.4 |
| 5 | 1.271 | 0.063 | 5.0 | 1.372 | 0.127 | 9.3 |
| 6 | 0.664 | 0.032 | 4.8 | 0.773 | 0.076 | 9.9 |
| 7 | 1.360 | 0.083 | 6.1 | 1.522 | 0.104 | 6.8 |
| 8 | 1.009 | 0.075 | 7.4 | 1.111 | 0.099 | 8.9 |
| 9 | 0.932 | 0.037 | 3.9 | 0.997 | 0.070 | 7.0 |
| 10 | 0.705 | 0.019 | 2.7 | 0.783 | 0.048 | 6.2 |

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

| Inter-Lab (n = 14) | | | |
|--------------------|-------|-------|--------|
| Panel # | Mean | S.D. | % C.V. |
| 1 | 1.75 | 0.172 | 9.8 |
| 2 | 1.061 | 0.167 | 15.7 |
| 3 | 1.322 | 0.173 | 13.1 |
| 4 | 0.307 | 0.037 | 11.9 |
| 5 | 1.129 | 0.140 | 12.4 |
| 6 | 0.655 | 0.077 | 11.8 |

1.271

0.974

0.840

0.677

12.0

12.1

13.1

7.9

0.153

0.117

0.110

0.053

**Inter-Lab (n = 14)**

8

9

10

)

P

7 *

{9}------------------------------------------------

Image /page/9/Picture/1 description: The image shows the logo for the U.S. Department of Health and Human Services. The logo consists of a circular seal with the words "DEPARTMENT OF HEALTH & HUMAN SERVICES • USA" arranged around the perimeter. Inside the circle is an abstract symbol that resembles an eagle or bird-like figure with three stylized lines forming its body and wings.

Food and Drug Administration 9200 Corporate Boulevard Rockville MD 20850

JUN 9 1999

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

Re: K983842

> Trade Name: Biotest Anti-EBV Recombinant EA IgM 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 542 of the Act for devices under the Electronic Product Radiation Control provisions, or other Federal laws or regulations.

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

#### 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 Autman

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

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

Page ***_ of _***

- J10(k) Number (if known): 4983892

Device Name: _________________________________________________________________________________________________________________________________________________________________

Indications For Use:

The Biotest EA IgM is an enzyme immunoassay for the detection of IgM 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 IgG, 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) | | | |
|--------------------------------------------|-----------------------------------------------------------------|---------------------------------------------------------|---------|--|
| | <div> Woody Dubois </div> <div>(Division Sign Off)</div> | | | |
| | Division of Clinical Laboratory Devices | | | |
| | <table><tr><td>510(k) Number</td><td>K983842</td></tr></table> | 510(k) Number | K983842 | |
| 510(k) Number | K983842 | | | |
| Prescription Use X<br>(Per 21 CFR 801.109) | OR | Over-The-Counter Use ______<br>(Optional Format 1-2-96) | | |