iDart Lyme IgM ImmunoBlot Kit

{0} FDA U.S. FOOD &amp; DRUG ADMINISTRATION # 510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION DECISION SUMMARY ASSAY ONLY ## I Background Information: A 510(k) Number K242872 B Applicant ID-FISH Technology, Inc. C Proprietary and Established Names iDart Lyme IgM ImmunoBlot Kit D Regulatory Information | Product Code(s) | Classification | Regulation Section | Panel | | --- | --- | --- | --- | | LSR | Class II | 21 CFR 866.3830 - Treponema Pallidum Treponemal Test Reagents | MI - Microbiology | ## II Submission/Device Overview: A Purpose for Submission: To obtain a substantial equivalence determination for the iDart Lyme IgM ImmunoBlot Kit for qualitative detection of anti-Borrelia burgdorferi IgM antibodies in human serum. B Measurand: Anti-Borrelia burgdorferi IgM antibodies. C Type of Test: Immunoblot assay. ## III Intended Use/Indications for Use: A Intended Use(s): See Indications for Use below. Food and Drug Administration 10903 New Hampshire Avenue Silver Spring, MD 20993-0002 www.fda.gov {1} K242872 - Page 2 of 10 # B Indication(s) for Use: The iDart Lyme IgM ImmunoBlot Kit is an immunoblot assay intended for the in vitro qualitative detection of IgM antibodies to *Borrelia burgdorferi* in human serum. The iDart Lyme IgM ImmunoBlot Kit is intended to detect antibodies to Lyme Screen Antigen (LSA) and multiple other *B. burgdorferi* antigens following a modified two-tier test methodology. Positive results from the iDart Lyme IgM ImmunoBlot Kit are supportive evidence for the presence of antibodies and exposure to *B. burgdorferi*. Negative results do not preclude infection with *B. burgdorferi*. iDart Lyme IgM ImmunoBlot Kit is intended to aid in the diagnosis of Lyme disease and the test kit should only be used on samples from patients with clinical history, signs and symptoms consistent with Lyme disease. The iDart Lyme IgM Immunoblot Kit is not intended as a screen for asymptomatic patients. Test results are to be used in conjunction with information obtained from the patient's clinical evaluation and other diagnostic procedures. # C Special Conditions for Use Statement(s): Rx - For Prescription Use Only # D Special Instrument Requirements: Not Applicable # IV Device/System Characteristics: # A Device Description: The iDart Lyme IgM ImmunoBlot is a line immunoblot assay. Antigenic proteins specific for different *Borrelia* species that cause Lyme Disease are produced by recombinant DNA technology in *Escherichia coli*. The purified proteins are applied as discrete lines on a nitrocellulose membrane (test strip) along with two control proteins in the following order: Control 1 (C1: IgG/IgM), Control 2 (C2: Protein L), P93, P41, P39, P23, P31, P34, and LSA (a chimeric VlsE peptide termed the Lyme Screening Antigen).  Figure 1. Schematic of the iDart Lyme IgM ImmunoBlot strip. # B Principle of Operation: During the test procedure, human serum is added to the test strip. Antibodies to *Borrelia burgdorferi*, if present, will bind to antigen lines on the test strip. After removing serum and unbound antibodies by washing, the test strip is incubated with alkaline phosphatase conjugated anti-human IgM antibody for detection of anti-*Borrelia burgdorferi* IgM antibodies in the patient sample. After removing the alkaline phosphatase conjugated antibody by washing, the antigen-antibody complex is visualized as bands on the test strip by adding the alkaline phosphatase substrate 5- {2} Bromo, 4-chloro, 3-indolylphosphate (BCIP) and nitro blue tetrazolium (NBT) to form a blue-purple precipitate on the detected band(s). The reaction stops by washing the test strip with distilled or deionized water. A test strip reading guide included with the kit shows the location of specific antigens in the test strip. Every test strip has two functional control bands (C1 and C2). The test strip is only valid when both controls bands are visible after completion of the test. Any band found having a visual intensity equal to or greater than the C2 control band intensity is considered as a positive band. ## Reagent and Materials The following reagents are provided as part of the iDart Lyme IgM ImmunoBlot Kit. Table 1. iDart Lyme IgM ImmunoBlot Kit, 50 assays per kit. | Component | Packaging | Volume/ Quantity | Part No. | | --- | --- | --- | --- | | Lyme IgM ImmunoBlot strips | 50ml tube | 50 strips | LMIBS03 | | IB Sample diluent | 60ml bottle | 55ml | IBSD03 | | IB Wash Buffer | 60ml bottle | 60ml | IBWB03 | | Milk powder | 0.5ml microcentrifuge tube | 0.75g | Milk03 | | Lyme IgM IB Conjugate | 60ml bottle | 60ml | LMIBC03 | | IB Phosphatase Substrate | 60ml bottle | 60ml | IBPS03 | | LYME IgM IB Positive Control | 0.5ml microcentrifuge tube | 120μl | LMIBP03 | | LYME IgM IB Negative Control | 0.5ml microcentrifuge tube | 120μl | LMIBN03 | | LYME IgM IB Package Insert | Paper | 1 each | LMIBPI | | LYME IgM IB Reading Guide | Laminated paper | 1 each | LMIBRG | The following are materials and reagents required but not provided with the kit: i. ImmunoBlot Incubation Tray (may be purchased from ID-FISH Technologies, Inc.) ii. Pipettor 10, 200, and 1000μl iii. Platform Rocker ## Quality Controls Control material should be tested in accordance with the guidelines or requirements of local, state, and/or federal regulations or accrediting organizations. To monitor the assay, reagent performance and day-to-day variation, positive controls for anti-Borrelia antibodies along with a negative control must be tested with each run. 1. All reportable bands should be present on positive control strip. If any of the reportable bands are absent on positive control, the test must be repeated. 2. No test bands are present. If the negative control strip shows 2 or more reportable bands with intensity equal to or greater than C2 control band, the test must be repeated. 3. C1 and C2 control bands must show in every test strip. ## Interpretation of Results A test strip result is valid only if both internal controls bands (C1 and C2) are clearly visible, and the negative and positive serum controls results are comparable to the pre-established profiles. Within each strip, C2 is the benchmark calibrator for test bands. The intensity of the bands on the sample test strip is then scored by comparing the intensity of the reportable bands to the intensity of C2 band within the same strip. K242872 - Page 3 of 10 {3} Table 2. Scoring of Protein bands intensity. | Band Intensity | Indicated by | | --- | --- | | - | No band detected | | +/- = I | A mark on the strip, band intensity < C2 control band | | + | A definite line or band intensity ≥ to C2 control band | Table 3. Interpretation of results for iDart Lyme IgM ImmunoBlot test. | Test Result | Interpretation | | --- | --- | | Positive | Positive or indetermined LSA band AND one or more from at least TWO of the following groups are present –P41, P39, P23, P31, and P34; | | Negative | If the band pattern does not meet the positive criteria. | # V Substantial Equivalence Information: A Predicate Device Name(s): Viramed Borrela All-In-One ViraChip Test Kit B Predicate 510(k) Number(s): K220016 C Comparison with Predicate(s): | Device & Predicate Device(s): | K242872 | K220016 | | --- | --- | --- | | Device Trade Name | iDart Lyme IgM ImmunoBlot Kit | Viramed Borrelia All-In-One ViraChip Test Kit | | General Device Characteristic Similarities | | | | Intended Use/Indications For Use | The iDart Lyme IgM ImmunoBlot Kit is an immunoblot assay intended for the in vitro qualitative detection of IgM antibodies to Borrelia burgdorferi in human serum. The iDart Lyme IgM ImmunoBlot Kit is intended to detect antibodies to Lyme Screen Antigen (LSA) and multiple other B. burgdorferi antigens following a modified two-tier test methodology. Positive results from the iDart Lyme IgM ImmunoBlot Kit are supportive evidence for the presence of antibodies and exposure to B. burgdorferi. Negative results do not preclude infection with B. burgdorferi. iDart Lyme IgM ImmunoBlot Kit is intended to aid in the diagnosis of Lyme disease and the test kit should | The Viramed Biotech AG Borrelia All-In-One ViraChip is an in vitro qualitative microarray assay for the detection of IgM and IgG antibodies to Borrelia burgdorferi in human serum. The assay is intended for testing serum samples from symptomatic patients or those suspected of Lyme Disease. It is intended to detect antibodies to VlsE and multiple other B. burgdorferi antigens following a modified two-tier test methodology. Positive results from the Viramed Biotech AG Borrelia All-In-One ViraChip are supportive evidence for the presence of antibodies and exposure to B. burgdorferi, the causative agent for Lyme disease. Negative results do not preclude infection with B. burgdorferi. Test | K242872 - Page 4 of 10 {4} | | only be used on samples from patients with clinical history, signs and symptoms consistent with Lyme disease. The iDart Lyme IgM Immunoblot Kit is not intended as a screen for asymptomatic patients. Test results are to be used in conjunction with information obtained from the patient’s clinical evaluation and other diagnostic procedures. | results are to be used in conjunction with information obtained from the patient’s clinical evaluation and other diagnostic procedures as an aid in diagnosis of Lyme disease. The Viramed Biotech AG Borrelia All-In-One ViraChip Test must be used with a ViraChip Reader and the ViraChip Software. | | --- | --- | --- | | Sample Type | Same | Serum | | Controls | Same | Positive Control Serum, Negative Control Serum | | Assay Type | Same | Qualitative | | General Device Characteristic Differences | | | | Antibodies Detected | IgM | IgM and IgG | | Antigens | P41, P39, P23, P31, P34, and LSA (a chimeric VlsE peptide) | VlsE, 93 kD, 58 kD, 45kD, 39 kD, 30 kD, 23kD, 21 kD, 19 kD, 18kD, and 17 kD antigens of B. burgdorferi | | Assay Technology | ImmunoAssay | Antigen coated wells (Microarrays) | | Reagents | Sample diluent, Wash Buffer, Milk powder, Conjugate Buffer, Substrate solution | 10X Wash Buffer, Sample Buffer, Chromogen/Substrate Solution | | Sample volume | 20 μL neat serum in 1000 ml sample diluent | Samples diluted 1:76 and 100 μL added per well | | Instrumentation | Manual | Automated with ViraChip Reader | VI Standards/Guidance Documents Referenced: Establishing the Performance Characteristics of in Vitro Diagnostic Devices for the Detection of Antibodies to Borrelia burgdorferi. Guidance for Industry and Food and Drug Administration Staff VII Performance Characteristics (if/when applicable): A Analytical Performance: 1. Precision/Reproducibility: The reproducibility of the iDart Lyme IgM ImmunoBlot Kit was evaluated in a study that included six anti-Borrelia IgM human serum samples at different analyte levels: high positive, two moderate positive, one low positive, one high negative, and one negative samples. The study was conducted across three sites, each with two operators over five non-consecutive days. On each of the five days, each operator performed one run, and each run included three replicates per sample. This generated a total of 90 replicates per sample (3 sites x 2 operators x 3 replicates x 5 days). There was 100% agreement on all bands on all runs, days, and operators at all three sites. K242872 - Page 5 of 10 {5} Table 4. iDart Lyme IgM ImmunoBlot Kit Reproducibility Study Results (all sites) | Sample # | Sample Type | IgM | # of Samples (+) | Expected Result | % agreement with expected result | | --- | --- | --- | --- | --- | --- | | MA | High Positive | P | 90/90 | P | 100% | | MB | Moderate Positive | P | 90/90 | P | 100% | | MC | Moderate Positive | P | 90/90 | P | 100% | | MD | Low Positive | P | 90/90 | P | 100% | | ME | High Negative | N | 0/90 | N | 100% | | MF | Negative | N | 0/90 | N | 100% | 2. Linearity: Not Applicable 3. Analytical Specificity/Interference: Cross-Reactivity: Potential cross-reactivity of the iDart Lyme IgM ImmunoBlot Kit was evaluated in a study that tested left-over patient sera containing antibodies to potentially cross-reacting conditions (e.g., viral and bacterial infections as well as autoimmune disorders). The table below summarizes the potential cross-reactant and the number of samples per cross-reactant included in the study. Cross-reactivity was observed with samples containing antibodies to *Leptospira* (2/10), mononucleosis (1/15), *H. pylori* (1/9), and parvovirus-19 (1/10). Table 5. Cross-Reactivity Study Results for iDart Lyme IgM ImmunoBlot Kit. | Source | Disease State | N (243) | IgM Positive | % Cross-reactivity | | --- | --- | --- | --- | --- | | CDC | Fibromyalgia | 15 | 0 | 0% | | | Mononucleosis | 15 | 1 | 6.67% | | | Multiple sclerosis | 15 | 0 | 0% | | | Rheumatoid arthritis | 11 | 0 | 0% | | | Severe periodontitis | 14 | 0 | 0% | | | Syphilis | 14 | 0 | 0% | | | Leptospira | 10 | 2* | 20.00%* | | IgeneX, Inc. | Rheumatoid Factor | 5 | 0 | 0% | | | ANA | 5 | 0 | 0% | | | Bartonella | 11 | 0 | 0% | | | Bartonella & Anaplasma | 1 | 0 | 0% | | | Bartonella &TBRF Borrelia | 1 | 0 | 0% | | | Babesiosis | 8 | 0 | 0% | | | Babesiosis & Tick-Borne Relapsing Fever | 2 | 0 | 0% | | | Babesiosis & Rickettsiosis | 1 | 0 | 0% | | | Tick Borne Relapsing Fever | 10 | 0 | 0% | | | Tick-Borne Relapsing Fever & Anaplasma | 1 | 0 | 0% | | | Tick-Borne Relapsing Fever & Ehrlichiosis | 1 | 0 | 0% | K242872 - Page 6 of 10 {6} | | Tick-Borne Relapsing Fever & Rickettsia | 1 | 0 | 0% | | --- | --- | --- | --- | --- | | | Anaplasmosis | 8 | 0 | 0% | | | Anaplasmosis & Ehrlichiosis | 1 | 0 | 0% | | | Ehrlichiosis | 4 | 0 | 0% | | | Rickettsiosis | 11 | 0 | 0% | | New York Biologics (NY) | HIV | 6 | 0 | 0% | | | HCV | 5 | 0 | 0% | | | HSV1 | 7 | 0 | 0% | | | CMV | 11 | 0 | 0% | | | EBV | 9 | 0 | 0% | | Kamineni Life Sciences Pvt. Ltd, | Pregnant women | 11 | 0 | 0% | | Hyderabad (India) | H. pylori | 9 | 1 | 11.11% | | Warde Medical Laboratory (MI) | Parvovirus-19 | 10 | 1 | 10.00% | | | Varicella-zoster virus | 10 | 0 | 0% | | False Positive | | | 5 | | | Agreement | | | 97.94% | | * Two Leptospira samples positive by iDart Lyme IgM testing were also positive for IgM with STTT. Endogenous Interference: The potential interfering effect of endogenous substances in patient samples when using the iDart Lyme IgM ImmunoBlot was evaluated using one high positive, one moderate positive, and one negative Borrelia IgM samples. Samples were spiked with the endogenous substances at the final concentrations listed in the table below. No interference effect was observed in the tested samples. Table 6. Endogenous Interference substances included in the study. | Endogenous Substance | Final Concentration | | --- | --- | | Bilirubin | 1 mg/dL | | | 15 mg/dL | | Albumin | 3.5 g/dL | | | 5 g/dL | | Cholesterol | 150 mg/dL | | | 250 mg/dL | | Triglycerides | 150 mg/dL | | | 500 mg/dL | | Hemoglobin | 10 g/dL | | | 20 g/dL | 4. Assay Reportable Range: Not Applicable 5. Traceability, Stability, Expected Values (Controls, Calibrators, or Methods): Fresh versus Frozen Sample Stability: This study was conducted to support the use of frozen samples in the clinical and analytical validation studies. To evaluate the performance of the iDart IgM ImmunoBlot Kit when using fresh and frozen samples, a total of 63 decoded left- K242872 - Page 7 of 10 {7} over patient serum samples were tested fresh (stored at 2° – 8°C) and after freezing at -20°C for at least 2 days, and not more than 22 days. All IgM positive samples (N=21) remained positive, and all IgM negative samples (N=42) remained negative when tested with the iDart Lyme IgM ImmunoBlot Kit fresh and after storage at -20°C. 6. Detection Limit: Not Applicable 7. Assay Cut-Off: Not Applicable ## B Comparison Studies: 1. Method Comparison with Predicate Device: The purpose of this study was to determine the clinical performance of the iDart Lyme IgM ImmunoBlot kit when compared to an FDA-cleared EIA and immunoblot for detection of antibodies to B. burgdorferi following as part of the standard two-tier test (STTT) methodology. All samples were tested using the iDart Lyme IgM ImmunoBlot Kit and the STTT comparator. A total of 997 serum samples were procured from a vendor and tested at three U.S. sites. Samples were collected in 2024 from different geographic regions in the U.S. All study samples were non-selected, left-over samples collected from patients with signs and symptoms consistent with Lyme disease that were prescribed a Lyme test. The table below summarizes the distribution of samples per testing site. Table 7: Sample distribution by clinical site and cohort. | Testing Site | Number of Samples | Sample Type | Vendors Providing Samples | | --- | --- | --- | --- | | Site 1 | 304 | Prospective | IGeneX Inc. | | Site 2 | 357 | Prospective | | | Site 3 | 336 | Prospective | | | Total | 997 | | | Additionally, the CDC Premarket panel containing 258 samples was also evaluated. This panel contains samples collected from patients diagnosed with Lyme Disease at different stages (Stages 1, 2, and 3), Lyme disease look-like infections (infectious mononucleosis, multiple sclerosis, rheumatoid arthritis, fibromyalgia and severe periodontitis), and from healthy controls living in both endemic and non-endemic regions of Lyme disease. ## Study Results ## Clinical samples All 997 samples were included in the performance calculations. The iDart Lyme IgM ImmunoBlot Kit Positive Percent Agreement (PPA) and Negative Percent Agreement (NPA) together with the 95% Confidence Interval (CI) were calculated against the comparator STTT. The table below summarizes the iDart Lyme IgM ImmunoBlot Kit results. K242872 - Page 8 of 10 {8} Table 8. iDart Lyme IgM ImmunoBlot Kit Performance (N=997) | | STTT | | | | --- | --- | --- | --- | | | | Positive (+) | Negative (-) | | iDart Lyme IgG ImmunoBlot | Positive (+) | 60 | 18 | | | Negative (-) | 6 | 913 | | | Total | 66 | 931 | | | PPA (95% CI) | 90.91% (81.55% – 95.77%) | | | | NPA (95% CI) | 98.07% (96.96% – 98.77%) | | CDC Premarket Panel The table below summarizes the results of the iDart Lyme IgM ImmunoBlot Kit when testing samples from the CDC Premarket panel. Table 9. CDC Premarket Panel: iDart Lyme IgM ImmunoBlot Kit Performance in Lyme Disease Samples. | Disease | Stage I | | Stage II | | Stage III | | Overall | | | --- | --- | --- | --- | --- | --- | --- | --- | --- | | N | 50 | | 9 | | 20 | | 79 | | | Test | iDart | STTT | iDart | STTT | iDart | STTT | iDart | STTT | | Positive | 33 | 28 | 8 | 7 | 11 | 9 | 52 | 44 | | Negative | 17 | 22 | 1 | 2 | 9 | 11 | 27 | 35 | | Agreement | 66.0% | 56.0% | 88.9% | 77.8% | 55.0% | 45.0% | 65.8% | 55.7% | Table 10: CDC Premarket Panel: iDart Lyme IgM ImmunoBlot Kit performance in control samples. | | Healthy Controls | | Disease Controls | | | --- | --- | --- | --- | --- | | N | 95 | | 84 | | | Test | iDart | STTT | iDart | STTT | | Positive | 2 | 1 | 1 | 3 | | Negative | 93 | 94 | 83 | 81 | | Agreement | 97.9% | 98.9% | 98.8% | 96.4% | 2. Matrix Comparison: Not Applicable C Clinical Studies: 1. Clinical Sensitivity: Not Applicable 2. Clinical Specificity: Not Applicable 3. Other Clinical Supportive Data (When 1. and 2. Are Not Applicable): Not Applicable D Clinical Cut-Off: Not Applicable K242872 - Page 9 of 10 {9} K242872 - Page 10 of 10 # E Expected Values/Reference Range: Well-characterized serum samples collected from apparent healthy individuals from both endemic and non-endemic areas for Lyme Disease were tested with the iDart Lyme IgM ImmunoBlot Kit following the instructions for use. Samples were provided by the CDC and the Bay Area Lyme foundation. Tables below summarize the performance of the device when testing samples from endemic and non-endemic areas respectively. One sample out of 177 (0.56%) tested samples from endemic areas was positive for IgM, using the iDart Lyme IgM ImmunoBlot Kit. Additionally, one sample out of 127 (0.79%) tested samples from non-endemic areas was positive for IgM using the iDart Lyme IgM ImmunoBlot Kit. Table 11: iDart Lyme IgM ImmunoBlot results for samples collected from healthy individuals in endemic areas. | Sample Source | N | IgM | % Positive | | --- | --- | --- | --- | | CDC | 50 | 1 | 2.00% | | Bay Area Lyme Foundation (NY, MA, WI) | 127 | 0 | 0.00% | | Total | 177 | 1 | 0.56% | | Agreement | | 99.44% | | Table 12: iDart Lyme IgM ImmunoBlot results for samples collected from healthy individuals in non-endemic areas. | Sample Source | N | IgM | % Positive | | --- | --- | --- | --- | | CDC | 45 | 1 | 2.22% | | IGeneX, Inc. | 82 | 0 | 0.00% | | Total | 127 | 1 | 0.79% | | Agreement | | 99.21% | | # VIII Proposed Labeling: The labeling supports the finding of substantial equivalence for this device. # IX Conclusion: The submitted information in this premarket notification is complete and supports a substantial equivalence decision.