BORRELIA BURGDORFERI IGM BLOT TEST

K113846 · Gold Standard Diagnostics · LSR · Jun 1, 2012 · Microbiology

Device Facts

• Record ID: K113846
• Device Name: BORRELIA BURGDORFERI IGM BLOT TEST
• Applicant: Gold Standard Diagnostics
• Product Code: LSR · Microbiology
• Decision Date: Jun 1, 2012
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 866.3830
• Device Class: Class 2

Intended Use

The Gold Standard Diagnostics Borrelia burgdorferi B31 IgM Line Blot Test Kit is intended for the qualitative detection of IgM antibodies to B. burgdorferi sensu stricto (B31) in human serum. This test is intended for use in testing human serum samples which have been found positive or equivocal using an ELISA or IFA test procedure to provide supportive evidence of infection with B. burgdorferi.

Device Story

The device is an in vitro diagnostic line blot assay for detecting IgM antibodies to B. burgdorferi. It utilizes nitrocellulose strips coated with purified B. burgdorferi antigens. Human serum samples are incubated on the strips; if specific IgM antibodies are present, they bind to the antigens. After washing, an enzyme-conjugated anti-human IgM is added, followed by a substrate (BCIP/NBT) that forms blue-violet precipitates at antigen-antibody binding sites. The resulting band pattern is visually interpreted by laboratory personnel to determine the presence or absence of specific IgM antibodies. The test is performed in a clinical laboratory setting as a secondary, supportive diagnostic step following initial screening by ELISA or IFA. The output provides qualitative evidence of infection, aiding clinicians in the diagnosis of Lyme disease.

Clinical Evidence

Prospective clinical study (n=310) compared subject device to predicate; 99.4% positive and negative percent agreement. Sensitivity evaluated on 100 clinically characterized samples (early, disseminated, late stages); subject device showed higher sensitivity than predicate across stages (e.g., 87.5% vs 77.5% in early stage). CDC reference panel testing showed high agreement with predicate. Analytical specificity 100% in 234 asymptomatic samples. Cross-reactivity tested against 215 specimens; minor cross-reactivity observed in Babesiosis, Ehrlichiosis, and Rickettsial disease samples, consistent with predicate performance.

Technological Characteristics

Line blot assay using purified or cloned B. burgdorferi proteins (23, 39, 41 kDa) sprayed onto nitrocellulose membranes. Detection via anti-human IgM antibody-enzyme conjugate and substrate-induced blue-violet precipitate. Manual interpretation of band patterns. No specialized instrumentation required.

Indications for Use

Indicated for qualitative detection of IgM antibodies to B. burgdorferi sensu stricto (B31) in human serum samples previously testing positive or equivocal by ELISA or IFA, to provide supportive evidence of Lyme disease infection.

Regulatory Classification

Identification

Treponema pallidum treponemal test reagents are devices that consist of the antigens, antisera and all control reagents (standardized reagents with which test results are compared) which are derived from treponemal sources and that are used in the fluorescent treponemal antibody absorption test (FTA-ABS), the Treponema pallidum immobilization test (T.P.I.), and other treponemal tests used to identify antibodies to Treponema pallidum directly from infecting treponemal organisms in serum. The identification aids in the diagnosis of syphilis caused by bacteria belonging to the genus Treponema and provides epidemiological information on syphilis.

Predicate Devices

• B. burgdorferi (IgM) Marblot Strip Test System (K951709)

Submission Summary (Full Text)

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# 2012

## 510(k) Summary

This 510(k) summary of safety and effectiveness information is being submitted in accordance with the requirement of SMDA 1990 and 21 CFR 807.92.

| 1) Submitter's Name: | Gold Standard Diagnostics |
|----------------------|------------------------------------|
| Address: | 2851 Spafford St. Davis, CA. 95618 |
| Phone Number: | 530-759-8000 |
| Contact Person: | Napoleon Monce |
| Date: | May 27, 2012 |

- 2) Product and Trade Name: Borrelia burgdorferi IgM B31 Line Blot Test Kit Common Name or Classification Name: Borrelia burgdorferi IgM Western blot
Product Code: LSR

### 3) Legally marketed device to which the submitter claims equivalence B. burgdorferi (IgM) Marblot Strip Test System (Distributed by Trinity Biotech) (K951709).

#### 4) Description of the device:

The assay requires a total of 70 minutes incubation time. The test uses purified antigens coated on nitrocellulose strips. Serum sample is added to each strip and incubated for 30 minutes. If Borrelia burgdorferi antibodies are present they will bind to the antigens on the strips. Unbound serum is removed by washing the wells three times. An enzyme-conjugated anti-human IgM is then added to each strip and incubated for 30 minutes. It will bind to the any antibody that was attached to the antigen on the strip. The wells are again washed three times followed by a DI water wash to remove any unbound conjugate. After unbound conjugate has been removed by a further washing step, a visualization of the antigenantibody-antibody complex is accomplished by the addition of a substrate which forms blue-violet precipitates at each site (antigen bands). The reaction is stopped by washing the nitrocellulose strip with distilled or deionized water. Depending on the observed band pattern one can interpret the presence or absence of specific IgM antibodies to B. burgdorferi infection.

#### 5) Intended use of the device

The Gold Standard Diagnostics Borrelia burgdorferi B31 IgM Line Blot Test Kit is intended for the qualitative detection of IgM antibodies to B. burgdorferi sensu stricto (B31) in human serum. This test is intended for use in testing human serum samples which have been found positive or equivocal using an ELISA or IFA test procedure to provide supportive evidence of infection with B. burgdorferi.

#### 6) Comparison with the predicate device

The Gold Standard Diagnostics Borrelia Burgdorferi B31 IgM Line Blot Test Kit was compared to a commercially marketed kit by Trinity Biotech the B. burgdorferi (IgM) Marblot Strip Test System

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(K951709). Both kits have the same intended use and use the same methodology. Below are tables comparing the reagents and the procedures of the two devices.

| Gold Standard Diagnostics B. burgdorferi<br>B31 IgM Line Blot Test Kit | Trinity Biotech B. burgdorferi (IgM)<br>Marblot Strip Test |
|------------------------------------------------------------------------|------------------------------------------------------------|
| Antigen coated nitrocellulose strips | Antigen coated nitrocellulose strips |
| IgM Conjugate - 100x | IgM Conjugate - 10x |
| Substrate – BCIP/NBT | Substrate |
| IgM Positive Control | IgM Band Locator Control |
| IgM Cutoff Control | IgM Weak Reactive Control |
| IgM Negative Control | IgM Negative Control . |
| Diluent/Wash Concentrate - 10x | Diluent/Wash Concentrate - 10x |

#### Table 1: Reagent Comparison

#### Table 2: Procedure Comparison

| Gold Standard Diagnostics B. burgdorferi<br><i>B31 IgM Line Blot Test Kit</i> | Trinity Biotech B. burgdorferi (IgM)<br>Marblot Strip Test |
|-------------------------------------------------------------------------------|------------------------------------------------------------|
| Dilute Samples 1:101 in Diluent/Wash | Dilute Samples 1:101 in Diluent/Wash |
| Add 15ul of Samples | Add 20ul of Samples |
| Incubate for 30 minutes while rocking | Incubate for 30 minutes while rocking |
| Wash three times with reconstituted | Wash three times with reconstituted |
| Diluent/Wash Solution | Diluent/Wash Solution |
| Add 1.5ml of reconstituted Conjugate | Add 2ml of reconstituted Conjugate |
| Incubate for 30 minutes while rocking | Incubate for 15 minutes while rocking |
| Wash three times with reconstituted | Wash three times with reconstituted |
| Diluent/Wash Solution | Diluent/Wash Solution |
| Wash one time with DI water | Wash one time with DI water |
| Add 1.5ml of Substrate | Add 2ml of Substrate |
| Incubate for 10 minutes ± 3 minutes while rocking | Incubate for 4-12 minutes while rocking |
| Wash three times with 1.5ml DI water | Wash three times with 2.0ml DI water |

#### 6(b1) Nonclinical Tests:

#### . Reproducibility

The reproducibility of the assay was done by testing a negative sample, a high negative sample, a low positive sample, and a moderate positive sample in triplicate for five days, twice a day, at three sites with two technicians per site giving a total of 90 data points per sample. Results of band reproducibility and sample reproducibility are shown below:

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#### Band Reproducibility

| Sample/kDa | 41 | 39 | 23 | Number<br>of Bands |
|----------------------|----|----|----|----------------------------|
| Negative | 0 | 0 | 0 | 0<br>significant<br>bands |
| High<br>Negative | 0 | 0 | 79 | ≤1<br>significant<br>bands |
| Low<br>Positive | 86 | 0 | 90 | 2<br>significant<br>bands |
| Moderate<br>Positive | 90 | 90 | 90 | 3<br>significant<br>bands |

#### Sample Reproducibility

| | | Final Interpretation | |
|-------------------|----------------------|----------------------|--------------|
| | Band Reproducibility | Positive | Negative |
| Sample | | | |
| Negative | 100% (0/0) | | 100% (90/90) |
| High Negative | 87.8% (79/90) | | 100% (90/90) |
| Low Positive* | 97.8% (176/180) | 95.6% (86/90) | |
| Moderate Positive | 100% (270/270) | 100% (90/90) | |

* A low positive sample is expected to yield a positivity of 95%.

#### Cross Reactivity

A cross reactivity study was performed on 215 specimens known to contain potentially cross reactive antibodies to B. burgdorferi. Sera from patients with infections and sera from patients with diagnoses that can be confused with the late manifestations of Lyme disease were tested. The results are summarized in the following table:

| Infection / Diagnosis | Number of sera tested | # Positive / (%) |
|------------------------------------------------------|-----------------------|------------------|
| Tick-borne relapsing fever /<br>Rickettsial Diseases | 23 | 1 / (4%) |
| Treponemal Infections | 12 | 0 / (0%) |
| Ehrlichiosis | 20 | 1 / (5%) |
| Babesiosis | 20 | 2 / (10%) |
| Leptospirosis | 1 | 0 / (0%) |

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| Parvovirus B19 | 9 | 0 / (0%) |
|------------------------|----|----------|
| Epstein-Barr Virus | 11 | 0 / (0%) |
| Cytomegalovirus | 32 | 0 / (0%) |
| <i>H. pylori</i> | 12 | 0 / (0%) |
| Fibromyalgia | 10 | 0 / (0%) |
| Rheumatoid Arthritis | 12 | 0 / (0%) |
| Herpes Simplex Virus | 16 | 0 / (0%) |
| Varicella Zoster Virus | 12 | 0 / (0%) |
| Autoimmune Disease | 25 | 0 / (0%) |

Two of the 20 Babesiosis samples and one each of the 23 Tick-borne relapsing fever / Rickettsial Disease and 20 Ehrlichiosis specimens were positive on the Gold Standard Diagnostics B. burgdorferi B31 IgM Line Blot Test. These samples were also tested on the predicate device which also gave positive results.

#### Interfering Substances

The effect of potential interfering substances on samples using the Gold Standard Diagnostics B. burgdorferi B31 IgM Line Blot Test was evaluated. High levels of hemoglobin, bilirubin, cholesterol and intralipids in serum samples were tested on six sera (two positive, one low positive, one high negative, and two negatives). The recommended concentrations from the guideline "Interference Testing in Clinical Chemistry" from the Clinical and Laboratory Standards Institute were used. The interferents at the concentrations tested did not have any influence on the band pattern. A small variability in band intensity was seen that is in the normal range of deviation and did not change the final interpretation. The results are summarized in the following table:

| Substance | Concentration | Interference |
|-------------|---------------|---------------|
| Hemoglobin | 2 g/l | None detected |
| Bilirubin | 342 µmol/l | None detected |
| Cholesterol | 13 mmol/l | None detected |
| Intralipids | 37 mmol/l | None detected |

#### Specimen Collection and Handling Conditions

A study was performed to substantiate the specimen storage and handling conditions for the Gold Standard Diagnostics B. burgdorferi B31 IgM Line Blot Test. Three samples were tested in triplicate after the following storage conditions:

- A) 2-8ºC for 24, 48, 72, 96, 120, 144 and 168 hours.
- B) Room temperature for 2, 6 and 8 hours.
- C) After 1, 5 and 10 freeze and thaw cycles.

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The samples gave consistent results for all testing parameters above, therefore the specimens can be held at 2-8°C for at least 168 hours or at room temperature for up to 8 hours and withstand up to 10 freeze and thaw cycles.

#### 8) Clinical Testing

#### Sensitivity Testing

A sensitivity study was performed on 100 clinically characterized samples obtained from Allen C. Steere, MD at the Massachusetts General Hospital. The samples encompassed early, disseminated, and late stages of Lyme disease. The results are summarized in the following table:

| | Number of<br>Samples | Line Blot Sensitivity<br>with 95% CI | Commercially Available<br>Device Sensitivity with<br>95% CI |
|--------------|----------------------|--------------------------------------|-------------------------------------------------------------|
| Early | 40 | 87.5% (35/40)<br>[73.2%-95.8%] | 77.5% (31/40)<br>[61.6%-89.2%] |
| Disseminated | 20 | 95.0% (19/20)<br>[75.1%-99.9%] | 85% (17/20)<br>[62.1%-96.8%] |
| Late | 40 | 27.5% (11/40)<br>[14.6%-43.9%] | 22.5% (9/40)<br>[10.8%-38.5%] |

#### Analytical Specificity (endemic & non-endemic)

For the determination of analytical specificity, testing of 234 asymptomatic samples from both endemic and non-endemic regions was performed. The results are summarized in the following table:

| Region | Number of Samples | Number Positive | Analytical Specificity |
|-------------|-------------------|-----------------|------------------------|
| Endemic | 115 | 0 | 100% |
| Non-endemic | 119 | 0 | 100% |

#### CDC Reference Panel

A standard panel of positive and negative specimens provided by the Center of Disease Control (CDC) for Lyme disease detection was tested both on the Gold Standard Diagnostics B. burgdorferi B31 IgM Line Blot Test and on the predicate device. The results are summarized in the following table:

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| Stage | Total | GSD Line Blot<br>% Agreement | Commercially<br>Available Device<br>% Agreement |
|-------------------------------|-------|------------------------------|-------------------------------------------------|
| Healthy | 5 | 100% (5/5) | 100% (5/5) |
| Early<br>(0-2 months) | 15 | 86.7% (13/15) | 73.3% (11/15) |
| Intermediate<br>(3-12 months) | 13 | 92.3% (12/13) | 84.6% (11/13) |
| Late<br>(>1year) | 7 | 57.1% (4/7) | 57.1% (4/7) |

#### Method Comparison

The performance of the Gold Standard Diagnostics B. burgdorferi B31 IgM Line Blot test was determined by conducting a prospective clinical study at three different geographic sites (Pennsylvania, North Carolina, and California) in the U.S. The patient samples were tested by an anti-B. burgdorferi total antibody ELISA test first and the resulting equivocal and positive specimens were tested on the Gold Standard Diagnostics B. burgdorferi B31 IgM Line Blot test and a commercially available B. burgdorferi IgM Blot test. The results are summarized in the following table:

| | | Trinity Biotech IgM Blot | |
|---------------------------|----------|--------------------------|----------|
| | | Positive | Negative |
| Gold Standard Diagnostics | Positive | | |
| IgM Line Blot | Negative | | |

Positive Percent Agreement = 99.4% (154/155) [95% CI: 96.5-100%] Negative Percent Agreement = 99.4% (154/155) [95% CI: 96.5-100%]

The discrepant samples were tested on a second commercially available assay. On the one Line Blot negative sample that was positive by the predicate, the second assay gave a positive result. Of the one Line Blot positive and predicate negative samples, the second assay called the sample positive.

The 310 samples were also tested by the Gold Standard B. burgdorferi IgG Line Blot test and 36.8% ( 14/310) were found to be positive and 63.2% (196/310) were found to be negative.

#### 9) Conclusion

From the performance data and kit comparison above, it is our conclusion that the Gold Standard Diagnostics B. burgdorferi B31 IgM Line Blot Test Kit is substantially equivalent to the B. burgdorferi (IgM) Marblot Strip Test System (K951709) commercially marketed by Trinity Biotech.

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#### DEPARTMENT OF HEALTH & HUMAN SERVICES

Image /page/6/Picture/1 description: The image shows the logo for the U.S. Department of Health & Human Services. The logo consists of a circular seal with the text "DEPARTMENT OF HEALTH & HUMAN SERVICES • USA" around the perimeter. Inside the circle is a stylized image of a human figure embracing a bird, which is a symbol representing the department's mission to protect and promote the health and well-being of Americans.

#### Food and Drug Administration

10903 New Hampshire Avenue Silver Spring, MD 20993

Gold Standard Diagnostics C/O Napoleon Monce Director, Product Development 2851 Spafford St. Davis, CA. 95618

JUN - 1 2012

Re: K113846

Trade/Device Name: Borrelia burgdorferi B31 IgM Line Blot Test Kit Regulation Number: 21 CFR 866.3830 Regulation Name: Treponema pallidum treponemal test reagents Regulatory Class: Class II Product Code: LSR Dated: December 28, 2011 Received: March 8, 2012

Dear Mr. Monce:

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.

If your device is classified (see above) into class II (Special Controls), it may be subject to such additional controls. Existing major regulations affecting your device can be found in Title 21, Code of Federal Regulations (CFR), Parts 800 to 895. 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 listing (21 CFR Part 807); labeling (21 CFR Parts 801 and 809); medical device reporting (reporting of medical device-related adverse events) (21 CFR 803); and good manufacturing practice requirements as set forth in the quality systems (QS) regulation (21 CFR Part 820). This letter

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#### Page 2 - Napoleon Monce

will allow you to begin marketing your device as described in your Section 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 Parts 801 and 809), please contact the Office of In Vitro Diagnostic Device Evaluation and Safety at [phone]. Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR 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/ReportaProblem/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 [phone] or [phone] or at its Internet address http://www.fda.gov/cdrh/industry/support/index.html.

Sincerely yours,

Uwe Schref for

Sally A. Hojvat, M.Sc., Ph.D. Director Division of Microbiology Devices Office of In Vitro Diagnostic Device Evaluation and Safety Center for Devices and Radiological Health

Enclosure

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# Indications for Use

510(k) Number (if known): K113846

Device Name: Borrelia burgdorferi B31 IgM Line Blot Test Kit

Indications For Use:

The Gold Standard Diagnostics Borrelia burgdorferi B31 IgM Line Blot Test Kit is intended for the qualitative detection of IgM antibodies to B. burgdorferi sensu stricto (B31) in human serum. This test is intended for use in testing human serum samples which have been found positive or equivocal using an ELISA or IFA test procedure to provide supportive evidence of infection with B. burgdorferi.

Prescription Use × (Part 21 CFR 801 Subpart D) AND/OR

Over-The-Counter Use (21 CFR 807 Subpart C)

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

Concurrence of CDRH, Office of In Vitro Diagnostic Devices (OIVD)

Tuare Felmberg

Division Sign-Off

Office of In Vitro Diagnostic Device Evaluation and Safety

510(K) 1/2 11 38 4 6

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