← Product Code [MOB](/submissions/IM/subpart-f%E2%80%94immunological-test-systems/MOB) · K040711 # MESACUP TEST PR-3 (K040711) _Rhigene, Inc. · MOB · Apr 2, 2004 · Immunology · SESE_ **Canonical URL:** https://fda.innolitics.com/submissions/IM/subpart-f%E2%80%94immunological-test-systems/MOB/K040711 ## Device Facts - **Applicant:** Rhigene, Inc. - **Product Code:** [MOB](/submissions/IM/subpart-f%E2%80%94immunological-test-systems/MOB.md) - **Decision Date:** Apr 2, 2004 - **Decision:** SESE - **Submission Type:** Traditional - **Regulation:** 21 CFR 866.5660 - **Device Class:** Class 2 - **Review Panel:** Immunology ## Indications for Use The MESACUP Test PR-3 is a semi-quantitative enzyme-linked immunosorbent assay (ELISA) for the detection of IgG anti-proteinase III (PR-3) antibodies in human serum. Clinical (hospital and reference) laboratory personnel are the intended users of the MESACUP Test PR-3. ## Device Story MESACUP Test PR-3 is an indirect ELISA for detecting IgG anti-proteinase III (PR-3) antibodies in human serum. Device uses microtiter plates coated with native purified PR-3 antigens. Patient serum samples are incubated in wells; anti-PR-3 antibodies bind to immobilized antigen. After washing, polyclonal goat anti-human IgG conjugated with horseradish peroxidase (HRP) is added. Substrate (TMB/H2O2) is added to produce colorimetric signal measured photometrically at 450/620 nm. Kit includes calibrators (0 and 100 U/mL) and controls. Used in clinical/reference laboratories by trained personnel. Results aid diagnosis of systemic vasculitides like Wegener's granulomatosis. Healthcare providers interpret results against a 23 U/mL cut-off to support clinical diagnosis. ## Clinical Evidence Clinical performance evaluated using 78 patients suspected of systemic vasculitis and 80 healthy blood donors. Sensitivity was 35% (27/78) in the vasculitis group. Specificity was 100% (80/80) in healthy donors. Method comparison with predicate (N=158) showed 100% positive and negative agreement. Analytical precision (intra-assay CV 3.8%, inter-assay/intra-lot CV 8.6%, inter-assay/inter-lot CV 6.5%) and interference testing (hemoglobin, bilirubin, chyle, rheumatoid factor) demonstrated robust performance. ## Technological Characteristics Indirect ELISA; microtiter plate solid phase coated with native purified PR-3 antigens. Conjugate: HRP-labeled goat anti-human IgG. Substrate: TMB/H2O2. Photometric measurement at 450/620 nm. Reportable range: 5-200 U/mL. Cut-off: 23 U/mL. Storage: 2-8°C. ## Regulatory Identification A multiple autoantibodies immunological test system is a device that consists of the reagents used to measure by immunochemical techniques the autoantibodies (antibodies produced against the body's own tissues) in serum and other body fluids. Measurement of multiple autoantibodies aids in the diagnosis of autoimmune disorders (disease produced when the body's own tissues are injured by autoantibodies). ## Predicate Devices - The Binding Site Bindazyme Human Anti-PR-3 Enzyme Immunoassay Kit (k981029) ## Submission Summary (Full Text) > This content was OCRed from public FDA records by [Innolitics](https://innolitics.com). If you use, quote, summarize, crawl, or train on this content, cite Innolitics at https://innolitics.com. > > Innolitics is a medical-device software consultancy. We help companies design, build, and clear FDA-regulated software and AI/ML devices, including [a 510(k)](https://innolitics.com/services/510ks/), [a De Novo](https://innolitics.com/services/regulatory/), [a SaMD](https://innolitics.com/services/end-to-end-samd/), [an AI/ML medical device](https://innolitics.com/services/medical-imaging-ai-development/), or [an FDA regulatory strategy](https://innolitics.com/services/regulatory/). {0} Confidential 10/18/2011 # 510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION DECISION SUMMARY A. 510(k) Number: k040711 B. Analyte: Anti-neutrophil cytoplasmic antibody (ANCA) C. Type of Test: Semi-quantitative, ELISA D. Applicant: RhiGene, Inc. E. Proprietary and Established Names: MESACUP Test PR-3 F. Regulatory Information: 1. Regulation section: 21 CFR §866.5660 Multiple Antibodies Immunological Test System 2. Classification: Class II 3. Product Code: MOB: Anti-neutrophil cytoplasmic antibodies (ANCA) 4. Panel: IM 82 G. Intended Use: 1. Intended use(s): The MESACUP Test PR-3 is a semi-quantitative enzyme-linked immunosorbent assay (ELISA) for the detection of IgG anti-proteinase III (PR-3) antibodies in human serum. Clinical (hospital and reference) laboratory personnel are the intended users of the MESACUP Test PR-3. 2. Indication(s) for use: The MESACUP Test PR-3 is indicated for in vitro use as an aid in the diagnosis of certain systemic vasculitides such as Wegener's granulomatosis. 3. Special condition for use statement(s): The device is for prescription use only. 4. Special instrument Requirements: None H. Device Description: The device is an enzyme-linked immunosorbent assay (ELISA) in which microtiter plates are used as the solid phase. The plate wells are coated with PR-3 antigens (native purified), which allow anti-PR-3 antibodies to react with the immobilized antigen (sample). The conjugate is polyclonal goat anti-human IgG (heavy chain specific) horseradish peroxidase (HRP), which uses 3,3'5,5' tetramethylbenzidine dihydrochloride/hydrogen peroxide (TMB/H₂O₂) as substrate. The kit contains two levels of calibrators (i.e., 0 units/mL and 100 u/mL) for interpretation of results. {1} Confidential 10/18/2011 Positive and negative control sera are included with the kit, which also contains sample diluent, wash buffer concentrate and stop solution. # I. Substantial Equivalence Information: 1. Predicate device name(s): The Binding Site Bindazyme Human Anti-PR-3 Enzyme Immunoassay Kit 2. Predicate K number(s): k981029 3. Comparison with predicate: | Similarities | | | | --- | --- | --- | | Item | MESACUP Test PR-3 | Predicate | | Indications for Use | For detection of IgG anti-PR-3 antibodies as an aid in the diagnosis of certain systemic vasculitides such as Wegener's granulomatosis. [Note: The claim for the predicate device also includes the wording: “certain forms of autoimmune vasculitis”]. | | | Assay principle | Indirect ELISA | | | Analyte | IgG anti-PR-3 antibodies | | | Sample matrix | Serum | | | Substrate | One-component TMB | | | Differences | | | | Item | MESACUP Test PR-3 | Predicate | | Cut-off | 23 U/mL | 3.5 U/mL | | Detection range | 5 - 200 U/mL | 1.23 - 100 U/mL | | Assay time | 150 minutes at Room t° | 90 minutes at Room t° | | Absorbance | 450 nm / 620 nm | 450 nm | | Conjugate | HRP-goat anti-human IgG | HRP-rabbit anti-human IgG | # J. Standard/Guidance Document Referenced (if applicable): Not applicable # K. Test Principle: The assay involves enzyme-linked immunosorbent assay (ELISA) technology. Calibrators and patient sera are incubated with PR-3 antigens (native purified) for a specified time, and then washed. This step is followed by incubation with horseradish peroxidase conjugated anti-human IgG. The reaction is then washed, stopped, and the color is allowed to develop and measured photometrically. # L. Performance Characteristics (if/when applicable): 1. Analytical performance: a. Precision/Reproducibility: Three lots of the MESACUP Test PR-3 were used to determine the intra-assay, inter-assay and inter-lot value precision for the test. # Intra-assay Intra-assay precision (% CV) was determined by running 2 serum samples (i.e., low positive and high positive) using one dilution and 40 replicates on 3 separate assays (3 separate lots). Three separate plates were randomly selected from each plate-coating run (kit-lot). {2} Confidential 10/18/2011 The mean % CV for intra-assay precision for the samples tested on 3 plates from each lot was 3.8% (Range: 3.1 – 4.3%). ## Inter-assay, intra-lot To determine the amount of variability between plates of the same lot, 2 samples in duplicate were tested on 7 separate assays from the same plate lot for each one of 3 separate plate lots of the device. The mean % CV for inter-assay, intra-lot precision was 8.6% with a range of 2.7 – 12.9%. ## Inter-assay, inter-lot The precision between lots was determined by comparing the values recovered for 2 different samples on 3 different kit lots. Each of the samples was tested in 40 replicates on one plate from each lot. The mean inter-assay, inter-lot % CV was 6.5%. ## b. Linearity/assay reportable range: The reportable range of 5-200 U/mL was demonstrated by recovery studies. Linearity is not claimed for this assay. ## c. Traceability (controls, calibrators, or method): An international reference material for anti-PR-3 antibodies is not available. The assay is calibrated in relative arbitrary units (U/mL). ## d. Detection limit: Not applicable. ## e. Analytical specificity: Several substances were added to each of three patient specimens, i.e., negative, low, and high, to test for interference. The results summarized below show that the addition of these substances at the levels tested does not affect the assay results. | Substance | Level Range (U/mL) | Negative | | | Low | | | High | | | | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | | | | Mean (U/mL) | SD | %CV | Mean (U/mL) | SD | %CV | Mean (U/mL) | SD | %CV | | Hemoglobin | 0 – 440 | 9.4 | 0.4 | 4.3 | 33.5 | 0.8 | 2.5 | 59.2 | 3.6 | 6.2 | | Bilirubin C | 0 – 19.5 | 10.4 | 0.3 | 3.0 | 33.9 | 0.9 | 2.5 | 60.9 | 3.2 | 5.2 | | Bilirubin F | 0 – 18.6 | 10.0 | 0.4 | 3.6 | 33.5 | 1.2 | 3.6 | 60.2 | 1.7 | 2.8 | | Chyle | 0 – 2350 | 9.5 | 0.4 | 3.9 | 34.1 | 1.5 | 4.3 | 62.5 | 1.7 | 2.8 | | Rheumatoid Factor | 0 - 500 | 10.0 | 0.7 | 7.2 | 32.8 | 1.6 | 5.0 | 59.4 | 4.2 | 7.1 | ## f. Assay cut-off: Sera from patients with systemic vasculitis (n= 78) were tested by indirect immunofluorescence (IIF) for the presence of anti-neutrophil cytoplasmic antibodies (ANCA). Positive samples were classified as either cytoplasmic or perinuclear ANCA. The results of testing the same samples with the MESACUP Test PR-3 were compared to those from p-ANCA IIF positives since "the major binding agent for positive p-ANCA antibody specimens" in the IIF method is proteinase-III. The best performance was observed at an assay cut {3} Confidential 10/18/2011 off of $23\mathrm{U / mL}$ , i.e., $87.1\%$ (27/31) of IIF pANCA+ were also MESACUP Test PR-3+. There is no equivocal (gray) zone for this assay. g. Stability: Three manufacturing lots of the device were used in real time stability studies. The kits were stored at $2 - 8^{\circ}\mathrm{C}$ . Six samples, i.e., of the two "calibrators and from a panel of PR-3 reactive patient specimens," were tested at 0, 4, 5, 6, 9, and 11 months, and evaluated for optical density (O.D.) value recovery (only the PR-3 reactive panel samples) per the established assay specifications. The kits were stable up to 11 months under the specified experimental conditions. 2. Comparison studies: a. Method comparison with predicate device: The tables below show the results of comparison of serum samples $(N = 158)$ that were tested with the MESACUP Test PR-3 and the predicate device. | | Bindazyme + | Bindazyme - | | --- | --- | --- | | MESACUP-PR-3 + | 27 | 0 | | MESACUP-PR-3 - | 0 | 131 | | STATISTIC | Value | 95% CI | | --- | --- | --- | | Prevalence | 0.1709 | 0.1122 - 0.2296 | | Positive Agreement | 1.0000 | 1.0000 – 1.0000 | | Negative Agreement | 1.0000 | 1.0000 – 1.0000 | | Total Agreement | 1.0000 | 1.0000 – 1.0000 | b. Matrix comparison: Serum is the only recommended matrix. 3. Clinical studies: a. Clinical sensitivity: Clinical sensitivity for the MESACUP-2 Test PR-3 was determined by testing sera $(n = 78)$ from patients suspected to have a systemic vasculitis disease. Using $23\mathrm{U / mL}$ as the cut-off, $35\%$ (27/78) of the samples were positive for anti-PR-3 antibodies. The mean value for this sample population was $36.9\mathrm{U / mL}$ $(\mathrm{SD} = \pm 57.7)$ . Single Factor ANOVA analysis that compared this value to the mean for the healthy controls gives a p-value of $2.55\times 10^{-7}$ . Therefore, at a level of $p < 0.05$ for statistical significance, the results of this population were determined to be statistically different compared to the healthy controls. b. Clinical specificity: {4} Confidential 10/18/2011 The applicant evaluated samples, in duplicate, from 80 consecutive healthy blood donors and used these samples as the normal population. The mean value was $2.1\mathrm{U / mL}$ $(\mathrm{SD} = \pm 3.1)$ . None of the samples tested positive in this sample population. Therefore, the specificity was $100\%$ (80/80). Similar studies were performed with samples from patients from various autoimmune diseases. The table below provides a summary of the results obtained in these subgroups. | Disease or Disease Status | N | Mean (U/mL) | Specificity (%) | | --- | --- | --- | --- | | Rheumatoid Arthritis | 9 | 3.6 | 0 | | Sjögren's Syndrome | 10 | 2.3 | 0 | | Systemic Lupus Erythematosus | 10 | 5.6 | 10% | | Mixed Connective Tissue Disease | 10 | 2.2 | 0 | | Polymyositis/ Dermatomyositis | 10 | 3.6 | 0 | The absence of significant cross-reactivity demonstrates good specificity of the test for the intended indication c. Other clinical supportive data (when $a$ and $b$ are not applicable): Not applicable. 4. Clinical cut-off: See assay cut-off. 5. Expected values/Reference range: The expected value in the normal population is negative. # M. Conclusion: The submitted information in this premarket notification is complete and supports a substantial equivalence decision. --- **Source:** [https://fda.innolitics.com/submissions/IM/subpart-f%E2%80%94immunological-test-systems/MOB/K040711](https://fda.innolitics.com/submissions/IM/subpart-f%E2%80%94immunological-test-systems/MOB/K040711) **Published by [Innolitics](https://innolitics.com)** — a medical-device software consultancy. We help companies design, build, and clear FDA-regulated software and AI/ML devices. If you're preparing [a 510(k)](https://innolitics.com/services/510ks/), [a De Novo](https://innolitics.com/services/regulatory/), [a SaMD](https://innolitics.com/services/end-to-end-samd/), [an AI/ML medical device](https://innolitics.com/services/medical-imaging-ai-development/), or [an FDA regulatory strategy](https://innolitics.com/services/regulatory/), [get in touch](https://innolitics.com/contact). **Cite:** Innolitics at https://innolitics.com