BORRELIA BURGDORFERI IGG BLOT TEST

{0} 1 # 510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION DECISION SUMMARY A. 510(k) Number: K113847 B. Purpose for Submission: New device C. Measurand: IgG antibodies to *Borrelia burgdorferi* proteins D. Type of Test: Western blot immunoassay E. Applicant: Gold Standard Diagnostics F. Proprietary and Established Names: *Borrelia burgdorferi* B31 IgG Line Blot Test Kit G. Regulatory Information: 1. Regulation section: 21 CFR 866.3830, Treponema pallidum treponemal test reagents 2. Classification: Class: II 3. Product code: LSR; Reagent, Borrelia Serological Reagent 4. Panel: 83 Microbiology {1} H. Intended Use: 1. Intended use(s): The Gold Standard Diagnostics Borrelia burgdorferi B31 IgG Line Blot Test Kit is intended for the qualitative detection of IgG 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. 2. Indication(s) for use: The Gold Standard Diagnostics Borrelia burgdorferi B31 IgG Line Blot Test Kit is intended for the qualitative detection of IgG 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. 3. Special conditions for use statement(s): For prescription use 4. Special instrument requirements: None I. Device Description: The Gold Standard Diagnostics Borrelia burgdorferi B31 IgG test is a line blot assay. The ten antigenic proteins specific for B. burgdorferi (sensu stricto) are either purified or cloned and expressed in the host E.coli. The purified individual proteins are transferred to a nitrocellulose membrane using a spraying micro-dispensing method. Positions of the lines are defined on the filter and the antigen bands are assigned in the following order: 18, 23, 28, 30, 39, 41, 45, 58, 66, and 93 kDa. During the test procedure, antibodies to Borrelia burgdorferi B31 (sensu stricto) present in the human serum sample will bind to the antigens coated onto the nitrocellulose strips. After removing serum and unbound antibodies by washing, the nitrocellulose strip is incubated with an antihuman IgG antibody-enzyme conjugate. After the unbound conjugate has been removed by a washing step, visualization of the antigen-antibody-antibody complex is accomplished by the addition of a substrate which forms a blue- {2} violet precipitate 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 IgG antibodies to $B$ . burgdorferi infection. # J. Substantial Equivalence Information: 1. Predicate device name(s): B. burgdorferi (IgG) Marblot Strip Test System from Trinity Biotech 2. Predicate 510(k) number(s): K950829 3. Comparison with predicate: | Similarities | | | | --- | --- | --- | | Item | Device | Predicate | | Intended use | The Gold Standard Diagnostics Borrelia burgdorferi B31 IgG Line Blot Test Kit is intended for the qualitative detection of IgG 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. | MarDx B. burgdorferi (IgG) Marblot Strip Test System is a Western blot assay for the qualitative in vitro detection of human IgG antibody to individual proteins of B. burgdorferi in human serum. The MarDx B. burgdorferi (IgG) Marblot Strip Test System is intended for use in testing human samples which have been found positive or equivocal using an EIA or IFA test procedure to provide supportive evidence of infection with B. burgdorferi. | | Specimen type | Serum | Serum | | Method | Qualitative | Qualitative | | Assay | Immunoblot | Immunoblot | {3} 4 | Differences | | | | --- | --- | --- | | Item | Device | Predicate | | Antigens | Individually purified or cloned proteins | Whole cell extract of *Borrelia burgdorferi* antigens | | Assay | Line blot | Western blot | K. Standard/Guidance Document Referenced (if applicable): Not applicable (N/A) L. Test Principle: Enzyme-immunoassay on a blot M. Performance Characteristics (if/when applicable): 1. Analytical performance: a. Precision/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 30 data points per sample. Results of band reproducibility and sample reproducibility are shown below: Band Reproducibility: | Sample/kDa | 93 | 66 | 58 | 45 | 41 | 39 | 30 | 28 | 23 | 18 | Number of Bands | | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | | Negative | | | | | 90 | | | | 90 | | <4 significant bands | | High Negative | 90 | | | 64 | 90 | | | | 90 | | 4 significant bands | | Low Positive | | | | 90 | 90 | 90 | | | 79 | 90 | 5 significant bands | | Moderate Positive | 90 | | 90 | | 90 | 90 | | 90 | | 90 | >5 significant bands | {4} 5 Sample Reproducibility: | | Band Reproducibility | Final Interpretation | | | --- | --- | --- | --- | | | | Positive | Negative | | Sample | | | | | Negative | 100% (180/180) | | 100% (90/90) | | High Negative | 92.8% (334/360) | | 100% (90/90) | | Low Positive | 97.6% (439/450) | 87.8%* (79/90) | | | Moderate Positive | 100% (540/540) | 100% (90/90) | | *A low positive sample is expected to yield a positivity of 95%. b. Linearity/assay reportable range: N/A c. Traceability, Stability, Expected values (controls, calibrators, or methods): N/A d. Detection limit: N/A e. Analytical specificity: Analytical Specificity Study: For the determination of the analytical specificity for the Gold Standard Diagnostics B. burgdorferi B31 IgG Line Blot Test, testing of 234 asymptomatic samples (blood donors) 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 | 2 | 98.6% | | Non-endemic | 119 | 0 | 100% | 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: {5} | Infection / Diagnosis | Number of Sera Tested | # Positive / (%) | | --- | --- | --- | | Tick-borne Relapsing Fever / Rickettsial Diseases | 23 | 1 / (4%) | | Treponemal Infections | 12 | 0 / (0%) | | Ehrlichiosis | 20 | 0 / (0%) | | Babesiosis | 20 | 2 / (10%) | | Leptospirosis | 1 | 0 / (0%) | | Parvovirus B19 | 9 | 0 / (0%) | | Epstein-Barr Virus | 11 | 0 / (0%) | | Cytomegalovirus | 32 | 0 / (0%) | | H. pylori | 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 of the 23 Tick-borne relapsing fever / Rickettsial Disease specimens were positive on the Gold Standard Diagnostics B. burgdorferi B31 IgG 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 IgG Line Blot Test was evaluated. High levels of hemoglobin, bilirubin, cholesterol and intralipids in serum samples were tested on six sera (two positives, 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: {6} | 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 | f. Assay cut-off: N/A ## 2. Comparison studies: a. Method comparison with predicate device: Method Comparison Study: The performance of the Gold Standard Diagnostics B. burgdorferi B31 IgG 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 ELISA test first and the resulting equivocal and positive specimens were tested on the Gold Standard Diagnostics B. burgdorferi B31 IgG Line Blot test and a commercially available B. burgdorferi IgG Blot test. The results are summarized in the following table: | | Predicate Device IgG Blot | | | | --- | --- | --- | --- | | | | Positive | Negative | | Gold Standard Diagnostics | Positive | 112 | 2 | | IgG Line Blot | Negative | 1 | 195 | Positive Percent Agreement = 99.1% (112/113) [95% CI: 95.2-100%] Negative Percent Agreement = 99.0% (195/197) [95% CI: 96.4-99.9%] 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 negative result. Of the two Line Blot positive and predicate negative samples, the second assay called one sample negative and the other sample positive. Sensitivity Study: A sensitivity study for the Gold Standard Diagnostics B. burgdorferi B31 IgG Line Blot Test was performed on 100 clinically characterized samples. The samples encompassed early, disseminated, and late stages of Lyme disease. The results are summarized in the following table: {7} 8 | Stage | Number of Samples | Line Blot Sensitivity with 95% CI | Predicate Device Sensitivity with 95% CI | | --- | --- | --- | --- | | Early | 40 | 7.5% (3/40) [1.6%-20.4%] | 7.5% (3/40) [1.6%-20.4%] | | Disseminated | 20 | 60.0% (12/20) [36.1%-80.9%] | 60.0% (12/20) [36.1%-80.9%] | | Late | 40 | 95.0% (38/40) [83.1%-99.4%] | 92.5% (37/40) [79.6%-98.4%] | CDC Reference Panel: A standard panel of positive and negative specimens provided by the Centers of Disease Control (CDC) for Lyme disease detection was tested both on the Gold Standard Diagnostics B. burgdorferi B31 IgG Line Blot Test and on the predicate device. The results are summarized in the following table: | Stage | Total | GSD Line Blot % Agreement | Predicate Device % Agreement | | --- | --- | --- | --- | | Healthy | 5 | 100% (5/5) | 100% (5/5) | | Early (0-2 months) | 15 | 93.3% (14/15) | 100% (15/15) | | Intermediate (3-12 months) | 13 | 84.6% (11/13) | 76.9% (10/13) | | Late (>1 year) | 7 | 100% (7/7) | 85.7% (6/7) | b. Matrix comparison: N/A 3. Clinical studies: a. Clinical Sensitivity: N/A b. Clinical specificity: N/A {8} c. Other clinical supportive data (when a. and b. are not applicable): N/A # 4. Clinical cut-off: # 5. Expected values/Reference range: The immune response to $B$ burgdorferi appears to follow a normal response pattern. IgM antibodies can be detected in some patients within days after onset, while IgG antibodies can be detected as early as two weeks after onset. IgM antibody titers often decrease several weeks to months after convalescence, but it is also possible that antibody titers remain constant up to several years. Band patterns will differ from sample to sample due to differences in patient immune responses and the stage to which the disease has progressed. The following table summarizes the frequency of bands observed with the Gold Standard Diagnostics $B$ burgdorferi B31 IgG Line Blot Test. | Stage / kDa | 93 | 66 | 58 | 45 | 41 | 39 | 30 | 28 | 23 | 18 | | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | | Early Lyme (n=55) | 5.5% | 7.3% | 20.0% | 20.0% | 80.0% | 16.4% | 10.9% | 1.8% | 45.5% | 27.3% | | Disseminated (n=33) | 27.3% | 27.3% | 45.5% | 45.5% | 84.8% | 54.5% | 9.1% | 21.2% | 63.6% | 45.5% | | Late Lyme (n=47) | 82.6% | 84.8% | 97.8% | 89.1% | 100% | 97.8% | 34.8% | 76.1% | 82.6% | 60.9% | | Prospective Samples (Not-staged) (n=310) | 12.6% | 30.3% | 16.8% | 35.5% | 48.4% | 36.5% | 19.7% | 8.6% | 36.1% | 38.7% | | Normal (n=115) (Non-Endemic) | 2.5% | 3.4% | 4.2% | 3.4% | 25.2% | 4.2% | 0.8% | 0.8% | 1.7% | 0.0% | | Normal (n=119) (Endemic) | 3.5% | 0.9% | 6.1% | 3.5% | 33.0% | 6.1% | 1.7% | 0.9% | 5.2% | 5.2% | {9} N. Proposed Labeling: 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. 10