AESKULISA DS DNA G
Device Facts
| Record ID | K041628 |
|---|---|
| Device Name | AESKULISA DS DNA G |
| Applicant | Aesku Diagnostics |
| Product Code | LSW · Immunology |
| Decision Date | Oct 14, 2004 |
| Decision | SESE |
| Submission Type | Traditional |
| Regulation | 21 CFR 866.5100 |
| Device Class | Class 2 |
Indications for Use
AESKULISA®dsDNA-G is a solid phase enzyme immunoassay with human recombinant double-stranded DNA (dsDNA) for the quantitative and qualitative detection of IgG antibodies against dsDNA in human serum. The assay is a tool in the diagnosis of systemic lupus erythematosus (SLE) and should be used in conjunction with other serological tests and clinical findings.
Device Story
AESKULISA®dsDNA-G is a solid-phase ELISA for detecting IgG antibodies against dsDNA in human serum. The device uses microtiter plates coated with human recombinant dsDNA. Patient serum is incubated; anti-dsDNA IgG antibodies bind to the antigen. Unbound components are washed away. Anti-human IgG HRP conjugate is added, followed by TMB substrate, producing a colorimetric reaction proportional to the antibody concentration. The reaction is stopped with 1M HCl and measured at 450 nm using a microplate reader. The device is used in clinical laboratories by trained personnel. Results aid in SLE diagnosis when interpreted alongside other serological tests and clinical findings. The assay provides quantitative and qualitative data to support clinical decision-making.
Clinical Evidence
No clinical data provided in the document; device clearance based on bench testing and performance validation of the immunoassay.
Technological Characteristics
Solid-phase enzyme immunoassay (ELISA). Utilizes human recombinant dsDNA as the capture antigen. Quantitative and qualitative detection of IgG antibodies. Manual or automated microplate processing. In vitro diagnostic use.
Indications for Use
Indicated for the quantitative and qualitative detection of IgG antibodies against dsDNA in human serum to aid in the diagnosis of systemic lupus erythematosus (SLE). For use in conjunction with other serological tests and clinical findings.
Regulatory Classification
Identification
An antinuclear antibody immunological test system is a device that consists of the reagents used to measure by immunochemical techniques the autoimmune antibodies in serum, other body fluids, and tissues that react with cellular nuclear constituents (molecules present in the nucleus of a cell, such as ribonucleic acid, deoxyribonucleic acid, or nuclear proteins). The measurements aid in the diagnosis of systemic lupus erythematosus (a multisystem autoimmune disease in which antibodies attack the victim's own tissues), hepatitis (a liver disease), rheumatoid arthritis, Sjögren's syndrome (arthritis with inflammation of the eye, eyelid, and salivary glands), and systemic sclerosis (chronic hardening and shrinking of many body tissues).
Related Devices
- K081251 — MODIFICATION TO AESKULISA DS DNA G · Aesku Diagnostics · May 13, 2008
- K974694 — IS-DSDNA TEST SYSTEM · Diamedix Corp. · Mar 3, 1998
- K994424 — DIAMEDIX IS-ANTI-DSDNA TEST SYSTEM · Diamedix Corp. · Feb 18, 2000
- K083381 — EUROIMMUN ANTI-DSDNA-NCX ELISA (IGG) · Euroimmun Us, Inc. · Apr 15, 2009
Submission Summary (Full Text)
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510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION
DECISION SUMMARY
A. 510(k) Number:
k041628
B. Purpose for Submission:
New device
C. Analyte:
Anti-DNA Antibodies
D. Type of Test:
ELISA, Qualitative and Quantitative
E. Applicant:
AESKU INC.
F. Proprietary and Established Names:
AESKULISA®dsDNA G Test
G. Regulatory Information:
1. Regulation section:
21 CFR § 866.5100, Antinuclear Antibody Immunological Test System
2. Classification:
Class II
3. Product Code:
LSW, Anti-DNA Antibody, Antigen and Control
4. Panel:
82 Immunology
H. Intended Use:
1. Intended use(s):
AESKULISA®dsDNA-G is a solid phase enzyme immunoassay with human recombinant double-stranded DNA (dsDNA) for the quantitative and qualitative detection of IgG antibodies against dsDNA in human serum. The assay is a tool in the diagnosis of systemic lupus erythematosus (SLE) and should be used in conjunction with other serological tests and clinical findings.
2. Indication(s) for use:
The assay is a tool in the diagnosis of systemic lupus erythematosus (SLE) and should be used in conjunction with other serological tests and clinical findings.
3. Special condition for use statement(s):
The device is for prescription use only.
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4. Special instrument Requirements:
Microtiter plate reader 450 nm reading filter and optional 620 nm reference filter (600-690 nm)
Microplate washing device (multichannel pipette or automated system)
I. Device Description:
AESKULISA®dsDNA-G is a solid phase enzyme immunoassay with human recombinant double-stranded DNA (dsDNA) for the quantitative and qualitative detection of IgG antibodies against dsDNA in human serum. The assay components include antigen coated microtiter plate, negative, positive and cut-off controls, wash buffer concentrate, sample buffer concentrate, anti-human IgG HRP horseradish peroxidase conjugate, 3,3',5,5' tetramethylbenzidine (TMB) substrate and 1M HCl stop solution. The positive and cut-off controls are composed of human serum of a specific dilution in standard buffer. The negative control contains standard buffer without any reactive component.
J. Substantial Equivalence Information:
1. Predicate device name(s):
Varelisa dsDNA ELISA
2. Predicate K number(s):
k930421
3. Comparison with predicate:
| Similarities | | |
| --- | --- | --- |
| Item | Device | Predicate |
| | AESKULISA dsDNA-G | Varelisa dsDNA ELISA |
| Intended Use | Quantitative and qualitative detection of IgG antibodies against dsDNA in human serum to aid in the diagnosis of SLE | Same |
| Assay Principle | Solid phase enzyme immunoassay | Same |
| Antigen | Recombinant plasmid DNA | Same |
| Differences | | |
| Item | Device | Predicate |
| Sample Matrix | Serum | Serum and plasma |
| Recommended normal ranges | Normal ≤15 IU/mL
Positive ≥15 IU/mL | Negative <35 IU/mL
Equivocal 35-55 IU/mL
Positive >55 IU/mL |
| Detection limit | 3.0 IU/mL | 1.0 IU/mL |
| Total incubation time | 60 minutes | 70 minutes |
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# K. Standard/Guidance Document Referenced (if applicable):
None referenced
# L. Test Principle:
Serum samples diluted 1:101 are incubated in the microplates coated with the specific antigen. Patient's antibodies, if present in the specimen, bind to the antigens. The unbound fraction is washed off in the following step. Afterwards anti-human immunoglobulins conjugated to horseradish peroxidase are incubated and react with the antigen-antibody complex of the samples in the microplates. Unbound conjugate is washed off in the following step. Addition of TMB substrate generates an enzymatic colorimetric (blue) reaction, which is stopped by diluted acid (color changes to yellow). The rate of color formation from the chromogen is a function of the amount of conjugate bound to the antigen-antibody complex and this is proportional to the initial concentration of the antibodies in the patient samples. The color formation is determined by measuring the OD value at $450\mathrm{nm}$ with a common photometer.
# M. Performance Characteristics (if/when applicable):
# 1. Analytical performance:
# a. Precision/Reproducibility:
Inter-assay - Inter-assay precision (%CV) was determined by running 3 different sera (low, medium, and high) 18 times on three different microtiter plates for five days. The results are as follows:
| Sample No. | Overall Mean (IU/mL) | Overall Mean %CV |
| --- | --- | --- |
| Low | 13.2 | 5.9 |
| Med | 151.6 | 2.3 |
| High | 463.3 | 2.6 |
Intra-assay - To determine intra-assay variation, 3 different sera (low, medium, high) were tested 24 times on 3 microtiter plates. The results are as follows:
| Sample No. | Overall Mean (IU/mL) | Overall Mean %CV |
| --- | --- | --- |
| Low | 12.71 | 5.91 |
| Med | 280.14 | 1.68 |
| High | 199.88 | 1.0 |
# b. Linearity/assay reportable range:
Linearity of the assay was demonstrated by doing serial dilutions of 6 sera from 1:100 to 1:1800. Linearity was set at 3-300 IU/mL
| Dilution .factor | dsDNA antibody Concentration (IU/mL) | | | | | |
| --- | --- | --- | --- | --- | --- | --- |
| | Serum 1 | Serum 2 | Serum 3 | Serum 4 | Serum 5 | Serum 6 |
| 1 | 331 | 73.4 | 775 | 153.1 | 42.9 | 179.4 |
| 0.5 | 139 | 31.1 | 375 | 71.18 | 21.6 | 86.4 |
| 0.25 | 48.16 | 12.32 | 120 | 28.87 | 10.8 | 41.8 |
| 0.125 | 20.24 | 5.73 | 43.37 | 12.18 | 5.4 | 19.8 |
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The regression analysis of the 6 sera gave the following results:
| Regression variables | Serum 1 | Serum 2 | Serum 3 | Serum 4 | Serum 5 | Serum 6 |
| --- | --- | --- | --- | --- | --- | --- |
| Intercept | -25.488 | -4.428 | -56.752 | -7.877 | 0.091 | -3.773 |
| Slope | 349.182 | 76.208 | 832.461 | 159.915 | 42.845 | 182.664 |
| r2 | 0.992 | 0.993 | 0.995 | 0.998 | 0.999 | 0.999 |
In addition, a recovery study was done at different dilution factors to see if the device will dilute linearly. However, due to the heterogeneous nature of human autoantibodies there might be samples that do not follow this rule. The results of this study are as follows:
| Sample No. | Dilution Factor | Measured conc IU/mL | Expected Conc IU/mL | % Recovery |
| --- | --- | --- | --- | --- |
| 1 | 1/100 | 42.9 | 43.2 | 99.3 |
| | 1/200 | 20.4 | 21.6 | 99.4 |
| | 1/400 | 9.3 | 10.8 | 86.1 |
| | 1/800 | 4.9 | 5.4 | 90.78 |
| 2 | 1/100 | 179.4 | 176.0 | 101.9 |
| | 1/200 | 86.4 | 88.0 | 98.2 |
| | 1/400 | 41.8 | 44.0 | 95.2 |
| | 1/800 | 19.8 | 22.0 | 90.0 |
c. Traceability (controls, calibrators, or method):
The AESKULISA®dsDNA-G calibrators are calibrated against the WHO/80 standard.
d. Detection limit:
The value for the detection limit was calculated as the mean of the optical densities (OD) of the standard buffer plus 3 times the standard deviation (SD), expressed in IU/mL. The mean plus 3 SD was $2.326\mathrm{IU / mL}$ . The detection limit was set at $3\mathrm{IU / mL}$ .
e. Analytical specificity:
Crossreactivity:
Ten sera from patients with various autoimmune diseases were tested on the AESKULISA®dsDNA-G and were found negative.
| disease | positive | negative |
| --- | --- | --- |
| CREST | 0 | 3 |
| Polymyositis | 0 | 1 |
| Scleroderma | 0 | 2 |
| MCTD | 0 | 1 |
| Wegener's granulomatosis | 0 | 1 |
| Sjogrens Syndrome | 0 | 1 |
| Sicca Syndrome | 0 | 1 |
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Interference:
Interference study was not performed. However, a statement not to use icteric, hemolyzed, or bacterially contaminated samples was included in the product insert.
f. Assay cut-off:
80 healthy donors were used for this study. The healthy donors were divided in the following age groups:
Young (<26y) 46
Middle age (26y-45y) 21
Elderly (46y+) 13
The mean plus 3 times the standard deviation was chosen as the cut-off value which was 15 IU/mL.
2. Comparison studies:
a. Method comparison with predicate device:
Fifty four SLE patient sera were tested on the AESKULISA®dsDNA-and the Varelisa® dsDNA.
| | Varelisa dsDNA | | | |
| --- | --- | --- | --- | --- |
| | | Pos | Neg | Total |
| AESKULISA dsDNA-2 | Pos | 34 | 13 | 47 |
| | Neg | 0 | 7 | 7 |
| | Total | 34 | 20 | 54 |
Negative agreement: 35.0%
Positive agreement: 72.3%
Total agreement 75.9%
The two devices gave conflicting results on 13 cases. All 13 cases are patients with SLE. The results are all positive on the AESKULISA but close to the cut-off value for the Varelisa. For this calculation, equivocal results for Varelisa were considered negative.
Because of the discrepancy in the method comparison with the predicate device, the sponsor was asked to make several dilutions of the WHO/80 standard and run them as unknowns on both devices. The results are as follows:
| WHO/80 dilution | Conc IU/mL | Varelisa dsDNA | | | AESKULISA dsDNA-2 | | |
| --- | --- | --- | --- | --- | --- | --- | --- |
| | | IU/mL | %CV | %recovery | IU/mL | %CV | %recovery |
| 1:200 | 100 | 125 | 4 | 125 | 98 | 7 | 98 |
| 1:400 | 50 | 55 | 4 | 110 | 48 | 1 | 96 |
| 1:800 | 25 | 29 | 2 | 115 | 28 | 8 | 112 |
| 1:1600 | 12.5 | 13 | 3 | 101 | 9 | 6 | 72 |
Twenty healthy donors on both devices and linear regression analysis showed (AESKULISA) = 1.15 (Varelisa) + 6.52 with r² =
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0.24. The correlation coefficient showed discrepancies at the low end but all healthy donors are below the respective cut-offs in both devices.
Twenty additional sera around the cut-off range of the devices were compared. The sera were from 19 female and 1 male, 2 belong to the group of <26 years; 10 to the group of 26-46 years and 8 to the group of 47+ years. A linear regression analyses was performed and the results are (AESKULISA) = 2.79 (Varelisa) - 18.75 with r² = 0.37
A histogram and a sigma plot with 95% confidence range were provided.
b. Matrix comparison:
Serum is the only recommended matrix.
3. Clinical studies:
a. Clinical sensitivity:
The AESKULISA®dsDNA-G detected 47 of 54 SLE patients who were positive for dsDNA antibodies. Clinical sensitivity is 87%.
b. Clinical specificity:
80 sera from healthy blood donors that were used in the cut-off study tested negative on the AESKULISA®dsDNA-G. Specificity was 100%
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 normal range for the assay is ≤ 15 IU/mL.
N. Conclusion:
The submitted material in this premarket notification is complete and supports a substantial equivalence decision.
