(265 days)
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No
The description details a standard chemiluminescent immunoassay run on an automated instrument. There is no mention of AI or ML in the device description, intended use, or performance studies. The quantitation relies on a predefined master curve and instrument-specific working curve, not on learning algorithms.
No.
Explanation: This device is an in vitro diagnostic (IVD) device used for the quantitative determination of IgG anti-double stranded deoxyribonucleic acid (dsDNA) antibodies in human serum to aid in the diagnosis of Systemic Lupus Erythematosus. It does not provide therapy or treatment.
Yes
Explanation: The "Intended Use / Indications for Use" section explicitly states, "The presence of anti-dsDNA antibodies, in conjunction with clinical findings and other laboratory tests, is an aid in the diagnosis of Systemic Lupus Erythematosus." This clearly indicates its role in aiding diagnosis, making it a diagnostic device.
No
The device is an immunoassay kit that includes physical reagents (paramagnetic beads, buffers, antibodies, calibrators, controls) and is designed to run on a specific hardware instrument (BIO-FLASH®). While the instrument includes software, the device itself is a combination of hardware and chemical components, not software-only.
Yes, this device is an IVD (In Vitro Diagnostic).
Here's why:
- Intended Use: The "Intended Use / Indications for Use" section explicitly states that the device is for the "quantitative determination of IgG anti-double stranded deoxyribonucleic acid (dsDNA) antibodies in human serum." It also states that the results are "an aid in the diagnosis of Systemic Lupus Erythematosus." This clearly indicates that the device is intended to be used in vitro (outside the body) to examine a human specimen (serum) for diagnostic purposes.
- Device Description: The description details a "chemiluminescent immunoassay" that analyzes "human serum" samples. This process involves chemical reactions and the measurement of light produced from these reactions, all performed on a sample taken from a patient.
- Components: The kit contains reagents, calibrators, and controls, which are typical components of an IVD assay used to perform tests on patient samples.
The definition of an IVD is a medical device intended to be used in vitro for the examination of specimens, including blood and tissue samples, derived from the human body, solely or principally for the purpose of providing information concerning a physiological or pathological state, or a congenital abnormality, or to determine the safety and compatibility with potential recipients, or to monitor therapeutic measures. This device fits this definition perfectly.
N/A
Intended Use / Indications for Use
QUANTA Flash dsDNA is a chemiluminescent immunoassay for the quantitative determination of IgG anti-double stranded deoxyribonucleic acid (dsDNA) antibodies in human serum. The presence of antidsDNA antibodies, in conjunction with clinical findings and other laboratory tests, is an aid in the diagnosis of Systemic Lupus Erythematosus.
QUANTA Flash dsDNA Calibrators are intended for use with the QUANTA Flash dsDNA chemiluminescent immunoassay for the determination of IgG anti-dsDNA antibodies in human serum. Each calibrator establishes a point of reference for the working curve that is used to calculate unit values.
QUANTA Flash dsDNA Controls are intended for use with the QUANTA Flash dsDNA chemiluminescent immunoassay for quality control in the determination of IgG anti-dsDNA antibodies in human serum.
Product codes (comma separated list FDA assigned to the subject device)
LSW, JIT, JJX
Device Description
QUANTA Flash dsDNA is a chemiluminescent microparticle immunoassay for the quantitative determination of IgG anti-double stranded deoxyribonucleic acid (dsDNA) antibodies in human serum. The QUANTA Flash dsDNA assay is designed to run on the BIO-FLASH® instrument. This platform is a fully automated closed system with continuous load and random access capabilities that automatically processes the samples, runs the assay and reports the results. It includes liquid handling hardware, luminometer and computer with software-user interface. The QUANTA Flash dsDNA assay utilizes a reagent cartridge format, which is compatible with the BIO-FLASH instrument.
Synthetic dsDNA is coated onto paramagnetic beads, which are stored in the reagent cartridge in suspension. When the assay cartridge is ready to be used for the first time, the entire cartridge is inverted several times to thoroughly mix the reagents. The sealed reagent tubes are then pierced with the reagent cartridge lid. The reagent cartridge is then loaded onto the BIO-FLASH instrument. Samples are also loaded onto the instrument in sample racks. A patient serum sample is prediluted 1:10 by the BIO-FLASH with system rinse in a disposable plastic cuvette. Small amounts of the diluted patient serum, the beads, and assay buffer are all combined into a second cuvette, and mixed. This cuvette is then incubated at 37°C. The beads are magnetized and washed several times. Isoluminol conjugated antihuman IgG antibodies are then added to the cuvette, and again incubated at 37°C. The beads are magnetized and washed repeatedly. The isoluminol conjugate is oxidized when Trigger 1 (Fe(III) coproporphyrin in sodium hydroxide solution) and Trigger 2 (urea-hydrogen peroxide in sodium chloride solution) are added to the cuvette, and the flash of light produced from this reaction is measured as Relative Light Units (RLU) by the BIO-FLASH optical system. The RLU are proportional to the amount of isoluminol conjugate that is bound to the human IgG, which is in turn proportional to the amount of anti-dsDNA antibodies bound to the corresponding dsDNA on the beads.
For quantitation, the QUANTA Flash dsDNA assay utilizes a predefined lot specific Master Curve that is uploaded onto the instrument through the reagent cartridge barcode. Every new lot number of reagent cartridge must be calibrated before first use, with the QUANTA Flash dsDNA Calibrators. The Master Curve is created during manufacturing by using in-house standards that are traceable to the First International Standard Preparation for dsDNA (WHO code: Wo/80). Based on the results obtained with the two Calibrators included in the Calibrator Set (sold separately), an instrument specific Working Curve is created, which is used to calculate international units (U)/mL from the instrument signal (RLU) obtained for each sample.
The QUANTA Flash dsDNA kit contains the following materials:
One (1) QUANTA Flash dsDNA Reagent Cartridge, containing the following reagents for 50 determinations:
- dsDNA antigen coated paramagnetic beads in a suspension.
- Assay Buffer 3 – buffer containing protein stabilizers and preservatives.
- Tracer IgG 2 Isoluminol labeled anti-human IgG antibodies in buffer, containing protein C. stabilizers and preservative.
The QUANTA Flash dsDNA Calibrators kit contains two vials of Calibrator 1 and two vials of Calibrator 2.
- QUANTA Flash dsDNA Calibrator 1: Two (2) barcode labeled tubes containing 0.7 mL prediluted, ready to use reagent. Calibrators contain human antibodies to dsDNA in stabilizers and preservatives.
- QUANTA Flash dsDNA Calibrator 2: Two (2) barcode labeled tubes containing 0.7 mL prediluted, ready to use reagent. Calibrators contain human antibodies to dsDNA in stabilizers and preservatives.
The QUANTA Flash dsDNA Controls kit contains two vials of Negative Control and two vials of Positive Control.
- QUANTA Flash dsDNA Low Control: Two (2) barcode labeled tubes containing 0.7 mL, ready to use reagent. Controls contain human antibodies to dsDNA in stabilizers and preservatives.
- QUANTA Flash dsDNA High Control: Two (2) barcode labeled tubes containing 0.7 mL, ready to use reagent. Controls contain human antibodies to dsDNA in stabilizers and preservatives.
Mentions image processing
Not Found
Mentions AI, DNN, or ML
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Input Imaging Modality
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Anatomical Site
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Indicated Patient Age Range
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Intended User / Care Setting
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Description of the training set, sample size, data source, and annotation protocol
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Description of the test set, sample size, data source, and annotation protocol
Description of the test set: Sample stability testing used thirteen samples encompassing negative, equivocal, and low to high positive samples. Cross-reactivity testing used 465 patient samples with various antibodies to autoimmune or infectious disease markers. Reference range establishment used a cohort of 171 subjects (121 apparently healthy blood donors, 7 viral hepatitis positive samples, 20 vasculitis, 19 rheumatoid arthritis, 4 HIV positive samples). Reference range verification used a second cohort of 210 samples (120 apparently healthy blood donors, 5 HBV positive samples, 5 HCV positive samples, 5 Syphilis, 5 HIV positive samples, 10 Sjogren's, 50 Rheumatoid arthritis, 10 anti-MPO positive). Clinical sensitivity and specificity were validated using a separate set of 1151 samples (20 Primary anti-phospholipid syndrome (PAPS), 50 Sjögren's syndrome (SS), 20 Celiac disease (CD), 40 Systemic sclerosis (SSc), 20 Idiopathic Inflammatory Myopathy (IIM), 20 MCTD, 20 Crohn's disease, 21 Graves Disease, 61 Hashimoto thyroiditis, 20 Hepatitis B, 18 Hepatitis C, 20 Syphilis, 101 Rheumatoid arthritis (RA), 34 Vasculitis, 20 Healthy controls, 22 Drug induced lupus (DIL), 644 Systemic lupus erythematosus (SLE)). Expected values were determined using a cohort of 300 apparently healthy blood donors (131 females and 169 males, ages 19 to 68 years). Method comparison with the predicate device used the same 1151 samples as the clinical validation study.
Summary of Performance Studies (study type, sample size, AUC, MRMC, standalone performance, key results)
Precision Study:
- Study Type: Evaluated according to CLSI EP5-A2.
- Sample Size: 9 samples.
- Key Results:
- Samples run in duplicates, twice a day, for 20 or 21 days.
- Total %CV values were within the acceptance limit (10%).
Reproducibility between sites (instruments) Study:
- Study Type: Performed in two sets. First set: 3 samples tested at 3 sites in quadruplicates, twice a day, for 10 days. Second set: 2 additional samples tested according to CLSI EP05-A3, at 3 sites in 5 replicates for 5 days.
- Sample Size: First set: 3 samples, 240 data points total per sample. Second Set: 2 samples, 75 data points total per sample.
- Key Results:
- All %CV values were within the acceptance limit (10%).
Reproducibility between lots Study:
- Study Type: Performed according to CLSI EP05-A3.
- Sample Size: 5 samples tested with 3 different lots of reagents in 5 replicates for 5 days, generating 75 data points total for each sample.
- Key Results:
- All %CV values were within the acceptance limit (10%).
Limit of Blank (LoB) and Limit of Detection (LoD) Study:
- Study Type: Consistent with CLSI EP17-A2 guideline.
- Sample Size: LoB: 4 aliquots of System Rinse tested in triplicates over 6 days on two reagent lots (72 data points per lot). LoD: 5 low level samples tested in 4 replicates for 3 days on two reagent lots (60 data points per lot).
- Key Results:
- LoD: 786 RLU (3 IU/mL).
- LoB: 520 RLU (2.2 IU/mL).
- Limit of Quantitation (LoQ): 1805 RLU (6.6 IU/mL).
Auto-rerun function and reportable results Study:
- Study Type: Validation study.
- Sample Size: Eight high positive specimens.
- Key Results:
- % recovery values for results obtained with auto-rerun compared to manual dilution were between 84% and 106% (average 96%), within the ± 20% acceptance limit.
Linearity Study:
- Study Type: Evaluated according to CLSI EPG-A.
- Sample Size: Five serum samples.
- Key Results:
- Percent recovery for all data points ranged from 90.8% to 113.0%.
- Slope (All Samples): 1.03 (0.98 to 1.05)
- R² (All Samples): 0.99 or higher for all samples.
Interference Study:
- Study Type: Performed according to CLSI EPO7-A2.
- Sample Size: Six specimens (24.3 IU/mL, 30.9 IU/mL, 37.9 IU/mL, 41.5 IU/mL, 132.3 IU/mL and 375.6 IU/mL) for bilirubin, hemoglobin, triglycerides, cholesterol. Six additional samples (21.4, 32.0, 40.3, 43.9, 140.3, and 364.0 IU/mL) for RF interference.
- Key Results:
- No interference detected with bilirubin up to 10 mg/dL (recovery: 86% to 102%), hemoglobin up to 200 mg/dL (recovery: 88% to 107%), triglycerides up to 1000 mg/dL (recovery: 91% to 109%), cholesterol up to 224.3 mg/dL (recovery: 91% to 109%), and RF IgM up to 947.1 IU/mL (recovery: 94% to 114%, or 3.6 to 3.7 IU/mL).
Cross-reactivity Study:
- Study Type: Testing for potential cross-reactivity.
- Sample Size: 465 patient samples with various antibodies to autoimmune or infectious disease markers.
- Key Results:
- 26 samples (5.6%) tested positive, proving the lack of cross-reactivity with autoimmune and infection induced antibodies.
Sample stability Study:
- Study Type: Stability testing.
- Sample Size: Thirteen samples.
- Key Results:
- All samples fulfilled the acceptance criteria (90-110% average recovery) for storage up to 17 days at 2-8°C, up to 48 hours at room temperature, and up to 6 freeze/thaw cycles.
Reagent stability (Shelf life) Study:
- Study Type: Accelerated stability studies and real-time stability studies.
- Key Results:
- Accelerated stability studies (4 weeks at 37 °C) supported a one-year expiration dating for components.
- Real-time stability data was provided for Calibrators up to 16 months, Controls up to 16 months, and reagent cartridge up to 21 months, and all results were within acceptance limits.
In-use (onboard) stability Study (Calibrators):
- Study Type: Onboard stability assessment.
- Key Results:
- 5 successful calibrations performed over an 8.5 hour period. Calibrator RLU values remained within the 90-110% range. All characterized patient samples ran within their expected range. Supports claim of up to 4 calibrations over an 8 hour period.
In-use (onboard) stability Study (Controls):
- Study Type: Onboard stability assessment.
- Key Results:
- Low and High Controls ran within their respective acceptable range for all 20 runs, with %CV values of 5.2 % and 7.7 % respectively.
- Linear regression line was within 85 % and 115 % at run 15 for both Controls.
In-use (onboard) stability Study (Reagent Cartridge):
- Study Type: In-use stability determination.
- Sample Size: Four serum specimens with different reactivity levels, Low and High Controls.
- Key Results:
- In-use (onboard) stability of the dsDNA reagent cartridge was set at 60 days.
Cut-off (reference range) establishment and verification Study:
- Study Type: Cut-off establishment and verification.
- Sample Size: Establishment: 171 subjects. Verification: 210 samples.
- Key Results:
- Cut-off: 27 IU/mL. Indeterminate range: 27-35 IU/mL. > 35 IU/mL is positive.
- Verification cohort showed 94.8% specificity.
Clinical sensitivity, specificity Study:
- Study Type: Validation study.
- Sample Size: 1151 samples (excluding drug induced lupus group and healthy controls for calculations - N=1109).
- Key Results:
- Indeterminate = Negative:
- Sensitivity: 42.7 % (38.9-46.6%)
- Specificity: 94.4 % (91.9 - 96.3%)
- Indeterminate = Positive:
- Sensitivity: 49.1 % (45.2-52.9%)
- Specificity: 87.5 % (84.2-90.2%)
- Sensitivity in lupus nephritis (N=25): 80% (20/25) for both indeterminate as negative and indeterminate as positive.
- Indeterminate = Negative:
Comparison with the predicate device Study:
- Study Type: Method comparison.
- Sample Size: All 1151 samples from the Validation Set study. Specifically, 481 samples were within the AMR of both devices. 333 samples were below the AMR of QUANTA Flash. 30 samples were above the AMR of QUANTA Flash. 124 samples were around the cut-off (25-40 IU/mL) with QUANTA Flash.
- Key Results:
- Within AMR (N=481), Indeterminate = negative:
- Positive Agreement: 80.3% (74.6 – 84.9%)
- Negative Agreement: 79.1% (73.6 - 83.6%)
- Total Agreement: 79.6% (75.8 – 83.0%)
- Within AMR (N=481), Indeterminate = positive:
- Positive Agreement: 68.5% (63.6 – 73.0%)
- Negative Agreement: 70.0% (60.9 – 77.8%)
- Total Agreement: 68.8% (64.5 – 72.8%)
- Samples below AMR (N=333): 98.8% agreement.
- Samples above AMR (N=30): 100% agreement.
- Samples around cut-off (N=124), Indeterminate = negative:
- Positive Agreement: 21.4% (7.6 – 47.6%)
- Negative Agreement: 75.5% (66.6 – 82.5%)
- Total Agreement: 69.4% (60.8 – 76.8%)
- Samples around cut-off (N=124), Indeterminate = positive:
- Positive Agreement: 90.0% (79.9 - 95.3%)
- Negative Agreement: 23.4% (14.7 - 35.1%)
- Total Agreement: 55.6% (46.9 - 64.1%)
- Within AMR (N=481), Indeterminate = negative:
Key Metrics (Sensitivity, Specificity, PPV, NPV, etc.)
Clinical sensitivity and specificity for SLE (N=1109):
- Indeterminate = Negative:
- Sensitivity: 42.7 % (38.9-46.6%)
- Specificity: 94.4 % (91.9 - 96.3%)
- Indeterminate = Positive:
- Sensitivity: 49.1 % (45.2-52.9%)
- Specificity: 87.5 % (84.2-90.2%)
- Sensitivity in lupus nephritis (N=25): 80% (20/25) for both indeterminate as negative and indeterminate as positive.
Agreement on samples within AMR (N=481):
- Indeterminate = negative:
- Pos. Agrmnt = 80.3% (74.6 – 84.9%)
- Neg. Agrmnt = 79.1% (73.6 - 83.6%)
- Total Agrmnt = 79.6% (75.8 – 83.0%)
- Indeterminate = positive:
- Pos. Agrmnt = 68.5% (63.6 – 73.0%)
- Neg. Agrmnt = 70.0% (60.9 – 77.8%)
- Total Agrmnt = 68.8% (64.5 – 72.8%)
Agreement on samples around the cut-off (25-40 IU/mL) (N=124):
- Indeterminate = negative:
- Pos. Agrmnt = 21.4% (7.6 – 47.6%)
- Neg. Agrmnt = 75.5% (66.6 – 82.5%)
- Total Agrmnt = 69.4% (60.8 – 76.8%)
- Indeterminate = positive:
- Pos. Agreement = 90.0% (79.9 - 95.3%)
- Neg. Agreement = 23.4% (14.7 - 35.1%)
- Total Agreement = 55.6% (46.9 - 64.1%)
Predicate Device(s): If the device was cleared using the 510(k) pathway, identify the Predicate Device(s) K/DEN number used to claim substantial equivalence and list them here in a comma separated list exactly as they appear in the text. List the primary predicate first in the list.
Reference Device(s): Identify the Reference Device(s) K/DEN number and list them here in a comma separated list exactly as they appear in the text.
Not Found
Predetermined Change Control Plan (PCCP) - All Relevant Information for the subject device only (e.g. presence / absence, what scope was granted / cleared under the PCCP, any restrictions, etc).
Not Found
§ 866.5100 Antinuclear antibody immunological test system.
(a)
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).(b)
Classification. Class II (performance standards).
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Food and Drug Administration 10903 New Hampshire Avenue Document Control Center - WO66-G609 Silver Spring, MD 20993-0002
April 11, 2016
INOVA Diagnostics, Inc. Dr. Gabriella Lakos Director, Assay Development 9900 Old Grove Road San Diego, CA 92131
Re: K152013
Trade/Device Name: QUANTA Flash® dsDNA OUANTA Flash® dsDNA Calibrators OUANTA Flash® dsDNA Controls Regulation Number: 21 CFR 866.5100 Regulation Name: Antinuclear Antibodies Immunological Test System Regulatory Class: II Product Code: LSW, JIT, JJX Dated: March 9, 2016 Received: March 11, 2016
Dear Dr. Lakos:
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. Please note: CDRH does not evaluate information related to contract liability warranties. We remind you, however, that device labeling must be truthful and not misleading.
If your device is classified (see above) into either class II (Special Controls) or class III (PMA), it may be subject to additional controls. Existing major regulations affecting your device can be found in the Code of Federal Regulations, Title 21, Parts 800 to 898. 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); good manufacturing practice requirements
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as set forth in the quality systems (QS) regulation (21 CFR Part 820); and if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR 1000-1050.
If you desire specific advice for your device on our labeling regulations (21 CFR Parts 801 and 809), please contact the Division of Industry and Consumer Education at its toll-free number (800) 638 2041 or (301) 796-7100 or at its Internet address
http://www.fda.gov/MedicalDevices/Resourcesfor You/Industry/default.htm. 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 Industry and Consumer Education at its toll-free number (800) 638-2041 or (301) 796-7100 or at its Internet address
http://www.fda.gov/MedicalDevices/ResourcesforYou/Industry/default.htm.
Sincerely yours,
Kelly Oliner -S
FOR
Leonthena R. Carrington, MS. MBA, MT(ASCP) Director Division of Immunology and Hematology Devices Office of In Vitro Diagnostics and Radiological Health Center for Devices and Radiological Health
Enclosure
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Indications for Use
510(k) Number (if known) K152013
Device Name QUANTA Flash® dsDNA QUANTA Flash® dsDNA Calibrators QUANTA Flash® dsDNA Controls
Indications for Use (Describe)
QUANTA Flash dsDNA is a chemiluminescent immunoassay for the quantitative determination of IgG anti-double stranded deoxyribonucleic acid (dsDNA) antibodies in human serum. The presence of anti-dsDNA antibodies, in conjunction with clinical findings and other laboratory tests, is an aid in the diagnosis of Systemic Lupus Erythematosus.
QUANTA Flash dsDNA Calibrators are intended for use with the QUANTA Flash dsDNA chemiluminescent immunoassay for the determination of IgG anti-dsDNA antibodies in human serum. Each calibrator establishes a point of reference for the working curve that is used to calculate unit values.
QUANTA Flash dsDNA Controls are intended for use with the OUANTA Flash dsDNA chemiluminescent immunoassay for quality control in the determination of IgG anti-dsDNA antibodies in human serum.
| Type of Use (Select one or both, as applicable) | |
|---|---|
| × Prescription Use (Part 21 CFR 801 Subpart D) | Over-The-Counter Use (21 CFR 801 Subpart C) |
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A Werfen Company
510(k) Summary
QUANTA Flash® dsDNA QUANTA Flash® dsDNA Calibrators QUANTA Flash® dsDNA Controls
Page 1 of 21
510(k) Summary
Table of Contents
| Administrative data |
|---|
| Device description |
| Intended use(s) |
| Substantial equivalence |
| Comparison to predicate device |
| Value assignment and traceability of Calibrators and Controls |
| Analytical performance characteristics |
| Precision |
| Reproducibility |
| Reproducibility between sites (instruments)…………………………………………………………………………………………………………………………………………………………………………………………………………………………………………………………………………………………………………………………………………………………… |
| Reproducibility between lots |
| Limit of Blank (LoB) and Limit of Detection (LoD) |
| Analytical Measuring Range (AMR) |
| Auto-rerun function and reportable results |
| High concentration hook effect |
| Linearity |
| Interference |
| Cross-reactivity |
| Sample stability |
| Reagent stability |
| Cut-off (reference range) establishment and verification |
| Clinical performance characteristics |
| Clinical sensitivity, specificity |
| Expected values |
| Comparison with the predicate device |
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This summary of the 510(k) safety and effectiveness information is being submitted in accordance with the requirements of SMDA 1990 and 21 CFR 807.92.
Administrative data
| Submitter: | Inova Diagnostics, Inc.
9900 Old Grove Road,
San Diego, CA, 92131 | |
|------------------------------|---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|----------------------------------------------------------------|
| Purpose of submission: | New device(s) | |
| Devices in the submission: | QUANTA Flash® dsDNA
QUANTA Flash® dsDNA Calibrators
QUANTA Flash® dsDNA Controls | |
| Scientific contact: | Gabriella Lakos, Director of Assay Development
Inova Diagnostics, Inc.
9900 Old Grove Road, San Diego, CA, 92131
Phone: 858-586-9900/1393
Fax: 858-863-0025
email: glakos@inovadx.com | |
| Quality Systems contact: | Ronda Elliott, VP of Quality Systems and Regulatory Affairs
Inova Diagnostics, Inc.
9900 Old Grove Road, San Diego, CA, 92131
Phone: 858-586-9900
Fax: 858-863-0025/1381
email: relliott@inovadx.com | |
| Preparation date: | 06/26/2015 | |
| Device name (assay kit): | Proprietary name: QUANTA Flash® dsDNA | |
| Common name: | Anti-dsDNA Chemiluminescent Immunoassay | |
| Classification name: | anti-dsDNA antibody, antigen and control | |
| Regulation Description | Antinuclear antibody immunological test system | |
| Regulation Medical Specialty | Immunology | |
| Review Panel | Immunology | |
| Product Code | LSW, Anti-DNA Antibody, Antigen and Control | |
| Regulation Number | 866.5100 | |
| Device Class | 2 | |
| Device name (Calibrators): | Proprietary name: | QUANTA Flash® dsDNA Calibrators |
| | Common name: | dsDNA Calibrators |
| | Classification name: | Calibrator, secondary |
| Regulation Description | Calibrator | |
| Regulation Medical Specialty | Clinical Chemistry | |
| Product Code | JIT | |
| Regulation Number | 862.1150 | |
| Device Class | 2 | |
| Device name (Controls): | Proprietary name: | QUANTA Flash® dsDNA Controls |
| | Common name: | dsDNA Controls |
| | Classification name: | Single (specified) analyte controls (assayed and
unassayed) |
| Regulation Description | Quality control material (assayed and unassayed) | |
| Regulation Medical Specialty | Clinical Chemistry | |
| Product Code | JJX | |
| Regulation Number | 862.1660 | |
| Device Class | 1 (reserved) | |
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Predicate device:
QUANTA Lite® dsDNA SC ELISA, 510(k) number: K993727
Device description
QUANTA Flash dsDNA is a chemiluminescent microparticle immunoassay for the quantitative determination of IgG anti-double stranded deoxyribonucleic acid (dsDNA) antibodies in human serum. The QUANTA Flash dsDNA assay is designed to run on the BIO-FLASH® instrument. This platform is a fully automated closed system with continuous load and random access capabilities that automatically processes the samples, runs the assay and reports the results. It includes liquid handling hardware, luminometer and computer with software-user interface. The QUANTA Flash dsDNA assay utilizes a reagent cartridge format, which is compatible with the BIO-FLASH instrument.
Synthetic dsDNA is coated onto paramagnetic beads, which are stored in the reagent cartridge in suspension. When the assay cartridge is ready to be used for the first time, the entire cartridge is inverted several times to thoroughly mix the reagents. The sealed reagent tubes are then pierced with the reagent cartridge lid. The reagent cartridge is then loaded onto the BIO-FLASH instrument. Samples are also loaded onto the instrument in sample racks. A patient serum sample is prediluted 1:10 by the BIO-FLASH with system rinse in a disposable plastic cuvette. Small amounts of the diluted patient serum, the beads, and assay buffer are all combined into a second cuvette, and mixed. This cuvette is then incubated at 37°C. The beads are magnetized and washed several times. Isoluminol conjugated antihuman IgG antibodies are then added to the cuvette, and again incubated at 37°C. The beads are magnetized and washed repeatedly. The isoluminol conjugate is oxidized when Trigger 1 (Fe(III) coproporphyrin in sodium hydroxide solution) and Trigger 2 (urea-hydrogen peroxide in sodium chloride solution) are added to the cuvette, and the flash of light produced from this reaction is measured as Relative Light Units (RLU) by the BIO-FLASH optical system. The RLU are proportional to the amount of isoluminol conjugate that is bound to the human IgG, which is in turn proportional to the amount of anti-dsDNA antibodies bound to the corresponding dsDNA on the beads.
For quantitation, the QUANTA Flash dsDNA assay utilizes a predefined lot specific Master Curve that is uploaded onto the instrument through the reagent cartridge barcode. Every new lot number of reagent cartridge must be calibrated before first use, with the QUANTA Flash dsDNA Calibrators. The Master Curve is created during manufacturing by using in-house standards that are traceable to the First International Standard Preparation for dsDNA (WHO code: Wo/80). Based on the results obtained with the two Calibrators included in the Calibrator Set (sold separately), an instrument specific Working Curve is created, which is used to calculate international units (U)/mL from the instrument signal (RLU) obtained for each sample.
The QUANTA Flash dsDNA kit contains the following materials:
One (1) QUANTA Flash dsDNA Reagent Cartridge, containing the following reagents for 50 determinations:
- dsDNA antigen coated paramagnetic beads in a suspension. a.
- b. Assay Buffer 3 – buffer containing protein stabilizers and preservatives.
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- Tracer IgG 2 Isoluminol labeled anti-human IgG antibodies in buffer, containing protein C. stabilizers and preservative.
The QUANTA Flash dsDNA Calibrators kit contains two vials of Calibrator 1 and two vials of Calibrator 2.
- QUANTA Flash dsDNA Calibrator 1: Two (2) barcode labeled tubes containing 0.7 mL prediluted, ready to use reagent. Calibrators contain human antibodies to dsDNA in stabilizers and preservatives.
- । QUANTA Flash dsDNA Calibrator 2: Two (2) barcode labeled tubes containing 0.7 mL prediluted, ready to use reagent. Calibrators contain human antibodies to dsDNA in stabilizers and preservatives.
The QUANTA Flash dsDNA Controls kit contains two vials of Negative Control and two vials of Positive Control.
- -QUANTA Flash dsDNA Low Control: Two (2) barcode labeled tubes containing 0.7 mL, ready to use reagent. Controls contain human antibodies to dsDNA in stabilizers and preservatives.
- QUANTA Flash dsDNA High Control: Two (2) barcode labeled tubes containing 0.7 mL, ready to use reagent. Controls contain human antibodies to dsDNA in stabilizers and preservatives.
Intended use(s)
QUANTA Flash dsDNA is a chemiluminescent immunoassay for the quantitative determination of IgG anti-double stranded deoxyribonucleic acid (dsDNA) antibodies in human serum. The presence of antidsDNA antibodies, in conjunction with clinical findings and other laboratory tests, is an aid in the diagnosis of Systemic Lupus Erythematosus.
QUANTA Flash dsDNA Calibrators are intended for use with the QUANTA Flash dsDNA chemiluminescent immunoassay for the determination of IgG anti-dsDNA antibodies in human serum. Each calibrator establishes a point of reference for the working curve that is used to calculate unit values.
QUANTA Flash dsDNA Controls are intended for use with the QUANTA Flash dsDNA chemiluminescent immunoassay for quality control in the determination of IgG anti-dsDNA antibodies in human serum.
Substantial equivalence
The QUANTA Flash dsDNA, the QUANTA Flash dsDNA Calibrators and the QUANTA Flash dsDNA Controls have the same intended use and assay principle as the predicate device.
Comparison to predicate device
QUANTA Flash dsDNA reagent kit
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| Similarities | ||
|---|---|---|
| Item | QUANTA Flash dsDNA | Predicate Device |
| Intended use | QUANTA Flash dsDNA is a | |
| chemiluminescent immunoassay for | ||
| the quantitative determination of IgG | ||
| anti-double stranded deoxyribonucleic | ||
| acid (dsDNA) antibodies in human | ||
| serum. The presence of anti-dsDNA | ||
| antibodies, in conjunction with clinical | ||
| findings and other laboratory tests, is | ||
| an aid in the diagnosis of Systemic | ||
| Lupus Erythematosus (SLE). | This assay is intended for the in-vitro | |
| measurement of specific IgG | ||
| autoantibodies against double | ||
| stranded deoxyribonucleic acid | ||
| (dsDNA) present in human serum, as | ||
| an aid to the diagnosis of systemic | ||
| lupus erythematosus (SLE), in | ||
| conjunction with other clinical | ||
| findings. | ||
| Assay methodology | Solid phase (heterogenous) | |
| immunoassay | Solid phase (heterogenous) | |
| immunoassay | ||
| Traceability | Traceable to the First International | |
| Standard Preparation for dsDNA | ||
| (WHO code: Wo/80) | Traceable to the First International | |
| Standard Preparation for dsDNA (WHO | ||
| code: Wo/80) | ||
| Sample type | Serum | Serum |
| Shelf life | One year | One year |
| Differences | ||
|---|---|---|
| Item | QUANTA Flash dsDNA | Predicate Device |
| Detection/ | ||
| Operating principle | Chemiluminescent immunoassay | Enzyme-linked immunosorbent assay |
| Solid phase | Paramagnetic microparticles (beads) | 96-well plate |
| Antigen | Synthetic dsDNA | Calf thymus dsDNA |
| Conjugate | Isoluminol conjugated anti-human IgG | HRP conjugated anti-human IgG |
| Calibration | Lot specific Master Curve + two | |
| Calibrators (Sold separately) | Five lot specific calibrators | |
| (Included in the kit) | ||
| Interpretation | 35 IU/mL Positive result | 75 IU/mL Positive result |
| Measuring Range | AMR = 9.8 – 666.9 IU/mL | |
| Reportable range = up to 6669.0 | 12.3 – 1000.0 IU/mL |
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| IU/mL | ||
|---|---|---|
| -- | ------- | -- |
QUANTA Flash dsDNA Calibrators
| Item | QUANTA Flash dsDNA Calibrators | Predicate Device |
|---|---|---|
| Intended use | QUANTA Flash dsDNA Calibrators are | |
| intended for use with the QUANTA | ||
| Flash dsDNA chemiluminescent | ||
| immunoassay for the determination of | ||
| IgG anti-dsDNA antibodies in human | ||
| serum. Each calibrator establishes a | ||
| point of reference for the working | ||
| curve that is used to calculate unit | ||
| values. | No separate intended use; calibrators | |
| are part of the kit. | ||
| Analyte | Anti-dsDNA antibodies | Anti-dsDNA antibodies |
| Method | QUANTA Flash dsDNA | |
| chemiluminescent immunoassay | QUANTA Lite dsDNA SC ELISA | |
| Unit | IU/mL | IU/mL |
| Matrix | Human serum, stabilizers, and | |
| preservative | Human serum, stabilizers, and | |
| preservative | ||
| Physico-chemical | ||
| characteristics | Liquid, prediluted, ready to use | Liquid, prediluted, ready to use |
| Storage | 2-8 °C | 2-8 °C |
| Shelf life | One year | One year |
QUANTA Flash dsDNA Controls
| Item | QUANTA Flash dsDNA Controls | Predicate Device |
|---|---|---|
| Intended use | QUANTA Flash dsDNA Controls are | |
| intended for use with the QUANTA | ||
| Flash dsDNA chemiluminescent | ||
| immunoassay for quality control in the | ||
| determination of IgG anti-dsDNA | ||
| antibodies in human serum. | No separate intended use; controls are | |
| part of the kit. | ||
| Analyte | Anti-dsDNA antibodies | Anti-dsDNA antibodies |
| Method | QUANTA Flash dsDNA | |
| chemiluminescent immunoassay | QUANTA Lite dsDNA SC ELISA | |
| Unit | IU/mL | IU/mL |
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| Matrix | Human serum, stabilizers, and
preservative | Human serum, stabilizers, and
preservative |
|-------------------------------------|-----------------------------------------------|-----------------------------------------------|
| Physico-chemical
characteristics | Liquid, ready to use | Liquid, prediluted, ready to use |
| Levels | 2 (low and high) | 2 (negative and positive) |
| Storage | 2-8 °C | 2-8 °C |
| Shelf life | One year | One year |
Value assignment and traceability of Calibrators and Controls
The QUANTA Flash dsDNA Calibrators and Controls are manufactured by diluting human serum that contains high titer of anti-dsDNA antibodies with antibody stabilizer buffer, containing preservative. The human serum is obtained from commercial sources and it is tested for markers of infectious substances.
The target IU/mL is achieved through trial dilutions on small scale. Once a dilution is selected, the Calibrators and Control are bulked, tested, and adjusted. Upon completion of the manufacturing process, the Calibrators and Controls are tested on at least two instruments, on at least two lots of reagent cartridge, in replicates of 10 to determine final value assignment.
The Master Curve Standards are traceable to the First International Standard Preparation for dsDNA (WHO code: Wo/80). Based upon this standardization, results are reported in International Units (IU)/mL. Calibrator and Control values are directly traceable to in-house Standards that are used to create the Master Curves for the QUANTA Flash dsDNA assay.
List of dsDNA Standards, Calibrators and Controls:
| Material | Assigned Value (IU/mL) |
|---|---|
| dsDNA Master Curve Standard 1 | 9.8 |
| dsDNA Master Curve Standard 2 | 19.4 |
| dsDNA Master Curve Standard 3 | 49.6 |
| dsDNA Master Curve Standard 4 | 90.7 |
| dsDNA Master Curve Standard 5 | 188.7 |
| dsDNA Master Curve Standard 6 | 375.6 |
| dsDNA Master Curve Standard 7 | 666.9 |
| Material | Manufacturing
Target Value (IU/mL) | Manufacturing
Target Range (IU/mL) |
|--------------------|---------------------------------------|---------------------------------------|
| dsDNA Calibrator 1 | 19 | 17 - 21 |
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| dsDNA Calibrator 2 | 189 | 169 - 209 |
|---|---|---|
| dsDNA Low Control | 22 | 19 - 25 |
| dsDNA High Control | 90 | 81 - 99 |
Analytical performance characteristics
Precision
The precision of the QUANTA Flash dsDNA assay was evaluated on 9 samples containing various concentrations of dsDNA antibodies in accordance with CLSI EP5-A2, Evaluation of Precision Performance of Quantitative Measurement Procedures - Approved Guideline: samples were run in duplicates, twice a day, for 20 or 21 days. Data were analyzed with the Analyse-it for Excel method evaluation software, and within run, between run, between day and total imprecisions are summarized in the Table below. Total %CV values were within the acceptance limit, 10%.
| QUANTA Flash® dsDNA | | Within-Run
(repeatability) | | Between-Run | | Between-Day | | Total | | |
|---------------------|----|-------------------------------|---------------|-------------|---------------|-------------|---------------|-----------|---------------|-----------|
| Sample
ID | N | Mean
(IU/mL) | SD
(IU/mL) | CV
(%) | SD
(IU/mL) | CV
(%) | SD
(IU/mL) | CV
(%) | SD
(IU/mL) | CV
(%) |
| 1 | 80 | 14.1 | 0.5 | 3.3 | 0.3 | 2.1 | 0.3 | 2.1 | 0.6 | 4.4 |
| 2 | 84 | 27.3 | 1.5 | 5.4 | 1.1 | 3.9 | 0.8 | 2.9 | 2.0 | 7.3 |
| 3 | 80 | 35.6 | 0.8 | 2.3 | 0.8 | 2.2 | 0.6 | 1.7 | 1.3 | 3.6 |
| 4 | 84 | 49.0 | 2.4 | 4.9 | 2.2 | 4.5 | 1.2 | 2.5 | 3.5 | 7.1 |
| 5 | 84 | 86.4 | 4.0 | 4.7 | 3.5 | 4.0 | 3.1 | 3.5 | 6.1 | 7.1 |
| 6 | 84 | 132.4 | 6.8 | 5.1 | 6.6 | 4.9 | 5.3 | 4.0 | 10.8 | 8.2 |
| 7 | 84 | 137.6 | 6.5 | 4.7 | 2.9 | 2.1 | 5.5 | 4.0 | 9.0 | 6.6 |
| 8 | 84 | 344.8 | 23.1 | 6.7 | 2.4 | 0.7 | 8.0 | 2.3 | 24.6 | 7.1 |
| 9 | 84 | 402.8 | 27.9 | 6.9 | 0.0 | 0.0 | 14.1 | 3.5 | 31.2 | 7.8 |
Reproducibility
Reproducibility between sites (instruments)
Studies were performed in two sets.
In the first set, three samples were tested at three different testing sites in quadruplicates, two times a day, for 10 days, to generate 80 data points per testing site, 240 data points total for each sample, using the same reagent lot.
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In the second set, two additional samples were tested according to CLSI EP05-A3 Evaluation of Precision of Quantitative Measurement Procedures, at three different testing sites in five replicates for 5 days, to generate 25 data points per testing site, 75 data points total for each sample, using the same reagent lot.
Data were analyzed with the Analyse-it for Excel method evaluation software, between sites imprecision was calculated, and the results are summarized in the Table below. All %CV values were within the acceptance limit, 10%.
| Sample ID | Mean (IU/mL) | N | Between Site Imprecision | |
|---|---|---|---|---|
| SD (IU/mL) | CV (%) | |||
| 1 | 32.6 | 240 | 2.0 | 6.1 |
| 2 | 52.9 | 240 | 1.3 | 2.5 |
| 3 | 142.7 | 240 | 3.9 | 2.8 |
| 4 | 13.2 | 75 | 0.0 | 0.0 |
| 5 | 466.1 | 75 | 24.8 | 5.3 |
Reproducibility between lots
Lot to lot reproducibility study was performed according to CLSI EP05-A3 Evaluation of Precision of Quantitative Measurement Procedures, by testing five samples with three different lots of reagents in five replicates for 5 days, to generate 25 data points per lot, 75 data points total for each sample.
Data were analyzed with the Analyse-it for Excel method evaluation software, between lots imprecision was calculated, and the results are summarized in the Table below. All %CV values were within the acceptance limit, 10%.
| Between Lot Imprecision | ||||
|---|---|---|---|---|
| Sample ID | Mean (IU/mL) | N | SD (IU/mL) | CV (%) |
| 1 | 13.9 | 75 | 0.7 | 5.2 |
| 2 | 25.7 | 75 | 0.2 | 0.7 |
| 3 | 33.1 | 75 | 2.5 | 7.5 |
| 4 | 85.9 | 75 | 4.5 | 5.3 |
| 5 | 349.3 | 75 | 24.7 | 7.1 |
Limit of Blank (LoB) and Limit of Detection (LoD)
The LoD of the QUANTA Flash dsDNA assay is 786 RLU (corresponding to 3 IU/mL. It was determined on two reagent lots, consistent with CLSI EP17-A2 guideline with proportions of false positives (alpha) less than 5% and false negatives (beta) less than 5%; based on 134 determinations, with 72 measurements on blank samples, and 60 measurements of low level samples, per reagent lot. The LoB is 520 RLU, , which corresponds to 2.2 IU/mL. The Limit of Quantitation (LoQ) is 1805 RLU (6.6 IU/mL). It was
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determined consistent with the quidelines in CLSI document EP17-A2, based on 120 determinations, and a Total Error goal of 20%. The LoB, the LoQ are below the AMR (below the lowest standard).
For the LoB study, 4 different aliquots of System Rinse were tested in triplicates over 6 days, on two reagent lots (72 data points per lot). The LoB was calculated separately on the two lots, and the higher value was used as the final LoB.
For the LoD study, five low level samples were used, with RLU values between the LoB and approximately 4 times of the LoB. Samples were tested in 4 replicates for 3 days on two reagent lots (60 data points per lot). The LoD was calculated separately on the higher value was used as the final LoD.
For the LoQ study, total imprecision was calculated for each sample that was used in the LoD study on each lot, and compared to the 20% Total Error goal. For each reagent lot, the sample with the lowest concentration that met accuracy specifications was taken as the LoQ for that lot. The greater LoQ across the lots was taken as the LoQ of the measurement procedure.
Analytical Measuring Range (AMR)
9.8 IU/mL - 666.9 IU/mL
The AMR is defined by the values of the lowest and highest Master Curve Standards.
Auto-rerun function and reportable results
The BIO-FLASH software has an Auto-rerun option available. If this option is selected, the instrument will automatically re-test any sample that has a result >666.9 IU/mL by further diluting it by a factor specified in the assay definition file (10 fold), thereby bringing the measured value within the AMR. The final result will be calculated by the software by taking into account the additional dilution factor. As the highest value that can be measured is 666.9 IU/mL, the highest value that can be reported is 6669.0 IU/mL.
To validate the Auto-rerun function, eight high positive specimens with results above the analytical measuring range were selected. The samples were run with the Auto-rerun function enabled on the BIO-FLASH. Then the specimens were manually diluted the same way as it happens in the Auto-rerun function (10 fold), and tested on the BIO-FLASH. The results were within the analytical measuring range after auto-rerun or manual dilution for all specimens. The % recovery values for results obtained with the auto-rerun results compared to the results obtained by manual dilution were between 84% and 106% (average 96%) and are within the ± 20% acceptance limit.
High concentration hook effect
Not applicable.
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Linearity
The linearity of the AMR was evaluated by a study according to CLSI EPG-A, Evaluation of the Linearity of Quantitative Measurement Procedures: A Statistical Approved Guideline. Five serum samples with various dsDNA antibody concentrations were diluted with stripped serum to obtain values that cover the AMR. Diluted samples were assayed in duplicate. Percent recovery was calculated compared to expected results (based on dilution). Percent recovery for all data points ranged from 90.8% to 113.0%. Obtained values were plotted against expected values, and linear regression analysis was performed on each samples, and also on the combined results. Acceptance criteria were 80%-120% (or ± 7 IU/mL) recovery, 0.9-1.1 slope and ≥0.95 R². Linear regression results are shown in the Table below.
| Sample ID | Test Range (IU/mL) | Slope (95% CI) | R2 |
|---|---|---|---|
| 1 | 20.0 to 200.2 | 1.00 (0.98 to 1.02) | 1.00 |
| 3 | 54.9 to 549.2 | 1.02 (0.98 to 1.06) | 0.99 |
| 4 | 71.6 to 644.5 | 1.08 (1.02 to 1.15) | 0.99 |
| 5 | 11.2 to 56.1 | 0.97 (0.93 to 1.01) | 0.99 |
| 7 | 65.1 to 585.8 | 1.00 (0.97 to 1.03) | 1.00 |
| All Samples | 11.2 to 644.5 | 1.03 (1.02 to 1.05) | 0.99 |
Interference
The interference study was performed according to CLSI EPO7-A2, Interference Testing in Clinical Chemistry; Approved Guideline - Second Edition. Six specimens were tested (24.3 IU/mL, 30.9 IU/mL, 37.9 IU/mL, 41.5 IU/mL, 132.3 IU/mL and 375.6 IU/mL). Interfering substances (hemoglobin, bilirubin, and triglycerides/cholesterol) were spiked into every specimen at three different concentrations in 10% of total specimen volume, and the resulting samples were assessed in triplicates with the QUANTA Flash dsDNA assay. Moreover, 6 additional samples (21.4, 32.0, 40.3, 43.9, 140.3, and 364.0 IU/mL) were tested for RF interference by combining them with different proportions of a high positive RF IgM serum sample (1894 IU/mL). Recovery of the unit values was calculated compared to control samples spiked with the same volume of diluents (10% of total sample volume, except for RF). For the RF interference study, recovery values were calculated compared to control samples created by adding negative serum to the test serum in the same proportions as the RF serum was used). Acceptance criteria for the interference studies were 85% - 115% recovery for positive samples, or ± 7 IU/mL difference for indeterminate and negative samples. The following interfering substances were tested:
| Interfering
substance | concentration #1
tested | concentration #2
tested | concentration #3
tested |
|--------------------------|----------------------------|----------------------------|----------------------------|
| Bilirubin, conjugated | 10 mg/dL | 5 mg/dL | 2.5 mg/dL |
| Hemoglobin | 200 mg/dL | 100 mg/dL | 50 mg/dL |
| Triglicerydes | 1000 mg/dL | 500 mg/dL | 250 mg/dL |
| Cholesterol | 224.3 mg/dL | 112.1 mg/mL | 56 mg/mL |
| RF IgM | 947.1 IU/mL | 568.2 IU/mL | 189.4 IU/mL |
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No interference was detected with bilirubin up to 10 mg/dL (recovery: 86% to 102%), hemoglobin up to 200 mg/dL (recovery: 88% to 107%), triglycerides up to 1000 mg/dL (recovery: 91% to 109%), cholesterol up to 224.3 mg/dL (recovery: 91% to 109%), and RF IgM up to 947.1 IU/mL (recovery: 94% to 114%, or 3.6 to 3.7 IU/mL).
Cross-reactivity
To test potential cross-reactivity with autoantibodies and infection-induced antibodies, 465 patient samples with various antibodies to autoimmune or infectious disease markers were tested. Altogether, 26 samples (5.6%) tested positive, proving the lack of cross-reactivity with autoimmune and infectioninduced antibodies:
| Patient Group | N | Positive | Indeterminate |
|---|---|---|---|
| Primary anti-phospholipid syndrome (PAPS) | 20 | 3 | 2 |
| Sjogren's syndrome (SS) | 50 | 3 | 1 |
| Celiac disease (CD) | 20 | 0 | 3 |
| Systemic sclerosis (SSc) | 40 | 3 | 4 |
| Idiopathic Inflammatory Myopathy (IIM) | 20 | 0 | 2 |
| MCTD | 20 | 3 | 3 |
| Crohn's disease | 20 | 1 | 0 |
| Graves Disease | 21 | 0 | 1 |
| Hashimoto thyroiditis | 61 | 3 | 4 |
| Hepatitis B | 20 | 0 | 1 |
| Hepatitis C | 18 | 0 | 1 |
| Syphilis | 20 | 1 | 0 |
| Rheumatoid arthritis (RA) | 101 | 8 | 6 |
| Vasculitis | 34 | 1 | 4 |
| Total | 465 | 26 | 32 |
| Percent | 5.6% | 6.9% |
Sample stability
Thirteen samples, encompassing negative, equivocal and low to high positive samples were tested in duplicates for up to 17 days at 2-8°C, up to 48 hours at room temperature, moreover, after repeated freeze/thaw cycles up to 6 cycles. Results were compared to those obtained on control samples (day zero, at 2-8°C)
Acceptance criteria: 90-110% average recovery.
All samples fulfilled the acceptance criteria at each time point for each condition.
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Based on these result, we recommend that samples are stored up to 48 hours at RT, up to 10 days of at 2-8 °C, and can be subjected to up to 3 freeze/thaw cycles (when samples are stored at or below -20 °C),
Reagent stability
Shelf life
Accelerated Stability
To establish the initial claim for shelf life, accelerated stability studies were performed for 4 weeks at 37 °C.
Accelerated stability testing was performed on three lots of each of the following sealed components of the QUANTA Flash dsDNA to establish initial stability claim:
- dsDNA beads
- Assay Buffer
- Tracer IgG
- Calibrators 1 and 2
- Low and High controls ●
Each week a new sealed component was placed in the incubator, and all components were tested at the end of the experiment together with the one that was stored at 5 ± 3°C. The recovery of the measured values was calculated for each time point (compared to those obtained with 5 ± 3°C stored reagent). All calculations were performed by comparing results of sealed components stored at 5 ± 3℃ (control) to those stored at 37 ± 3°C (test) for 1, 2, 3, and 4 weeks, where one week is equal to six months at 5 ± 3°C. Linear regression analysis was performed between recovery values and the number of days.
Acceptance criteria for one year preliminary expiration dating were:
-Microparticles (beads), Assay Buffer, and Tracer IgG:
With regression analysis, the 95% Cl interval of the regression line is between 85 and 115 % at 2 weeks, and no individual data point is outside the 75-125 %recovery range at 2 weeks.
- Controls and Calibrators:
With regression analysis, the lower 95% Cl interval of the regression line is between 90 and 110 % at 2 weeks, and no individual data point is outside the 80-120 %recovery range at 2 weeks.
All components tested fulfilled the acceptance criteria above, so one year expiration dating was assigned to each component.
Real time stability
To confirm the shelf life that was assigned based on accelerated stability studies, real time stability studies have been performed.
Real time stability testing was performed on Calibrators, Controls and reagent cartridge at regular time points to support the one year expiration.
We are providing real time stability data for Calibrators up to 16 months, Controls up to 16 months, and reagent cartridge up to 21 months.
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Controls were tested in triplicates at each time point.
- Acceptance criteria: results should fall within their acceptable ranges as it was established at the release of the controls.
Calibrators were tested with two different protocols, due to internal procedure change during the study. In the first part of the study Calibrators were used to calibrate a reagent cartridge and a panel of QC samples was tested.
- Acceptance criteria: results of the samples should fall within their respective QC ranges
In the second part of the study Calibrators were tested in triplicates at each time point as samples. Averages of the triplicates were compared to the value that was assigned to the Calibrators at release. - Acceptance criteria: % recovery of the average of the triplicates is between 85 and 115%, and %CV of the triplicates is 35 IU/mL are positive.
If a result falls in the indeterminate range, it means that the probability of SLE is low, but it is still higher than in the normal population. Forty-one out of the 644 SLE patients (6.4%) that were tested during the validation studies had their results fall in the indeterminate range. The frequency of indeterminate antidsDNA results in the tested normal population (n=300) was 2.0%. As several studies have documented a close relationship between antibody titer and disease activity, indeterminate anti-dsDNA results are more likely to occur in inactive disease compared to newly diagnosed SLE.
To verify the reference range, a second cohort of samples, consisting of 210 samples was tested. This cohort had similar composition as the first reference cohort:
| Diagnosis | Number | Number of equivocal | Number of positive |
|---|---|---|---|
| Apparently healthy blood donor | 120 | 1 | 2 |
| HBV positive samples | 5 | 0 | 1 |
| HCV positive samples | 5 | 0 | 0 |
| Syphilis | 5 | 0 | 1 |
| HIV positive samples | 5 | 0 | 0 |
| Sjogren's | 10 | 0 | 1 |
| Rheumatoid arthritis | 50 | 0 | 4 |
| anti-MPO positive | 10 | 0 | 0 |
| Total | 210 | 1 | 10 |
There were 11 samples above the cutoff in this cohort, resulting in 94.8% specificity. These results verify the reference range that was set based on the first cohort.
Clinical performance characteristics
Clinical sensitivity, specificity
A separate set of samples, none of which were used in establishing the reference range, was used to validate the clinical performance of the QUANTA Flash dsDNA. A total of 1151 samples were included in the Validation Set for the QUANTA Flash dsDNA.
Distribution of the cohort used in the QUANTA Flash dsDNA validation study:
| QUANTA Flash dsDNA, n pos | |||
|---|---|---|---|
| Patient Group | N | ind=pos | ind=neg |
| Primary anti-phospholipid syndrome (PAPS) | 20 | 5 | 3 |
| Sjögren's syndrome (SS) | 50 | 4 | 3 |
| Celiac disease (CD) | 20 | 3 | 0 |
| Systemic sclerosis (SSc) | 40 | 7 | 3 |
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| QUANTA Flash dsDNA, n pos | |||
|---|---|---|---|
| Patient Group | N | ind=pos | ind=neg |
| Idiopathic Inflammatory Myopathy (IIM) | 20 | 2 | 0 |
| MCTD | 20 | 6 | 3 |
| Crohn's disease | 20 | 1 | 1 |
| Graves Disease | 21 | 1 | 0 |
| Hashimoto thyroiditis | 61 | 7 | 3 |
| Hepatitis B | 20 | 1 | 0 |
| Hepatitis C | 18 | 1 | 0 |
| Syphilis | 20 | 1 | 1 |
| Rheumatoid arthritis (RA) | 101 | 14 | 8 |
| Vasculitis | 34 | 5 | 1 |
| Healthy controls * | 20 | 0 | 0 |
| Drug induced lupus (DIL) * | 22 | 2 | 2 |
| Total controls | 507 | 60 | 28 |
| Systemic lupus erythematosus (SLE) | 644 | 316 | 275 |
| Total | 1151 | 376 | 303 |
- Not included in the clinical sensitivity and specificity calculations
Clinical sensitivity and specificity for SLE is calculated in the tables below. Calculations were done in two ways; with equivocal results as negative. The drug induced lupus group and the healthy controls group were excluded from sensitivity and specificity calculations.
| Clinical Analysis (N=1109)
Indeterminate = Negative | Diagnosis | | | Analysis
(95% confidence) |
|--------------------------------------------------------|-----------|----------|-------|------------------------------------|
| | SLE | Controls | Total | |
| QUANTA Flash®
dsDNA | 275 | 26 | 301 | Sensitivity: 42.7 % (38.9-46.6%) |
| | 369 | 439 | 808 | Specificity: 94.4 % (91.9 - 96.3%) |
| | 644 | 465 | 1109 | |
| Clinical Analysis (N=1109) | Diagnosis | Analysis | ||||
|---|---|---|---|---|---|---|
| Indeterminate = Positive | SLE | Not SLE | Total | (95% confidence) | ||
| QUANTA Flash® | ||||||
| dsDNA | Positive | 316 | 58 | 374 | Sensitivity: 49.1 % (45.2-52.9%) | |
| Negative | 328 | 407 | 735 | Specificity: 87.5 % (84.2-90.2%) | ||
| Total | 664 | 465 | 1109 |
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For comparison, clinical sensitivity and specificity were calculated on the results obtained with the predicate device on the same population. The summary of the results is shown below:
| Clinical Performance (N=1109) | QUANTA Flash dsDNA | QUANTA Lite dsDNA SC |
|---|---|---|
| Indeterminate as positive | ||
| Sensitivity % (95% CI) | 49.1 (45.2-52.9) | 57.1 (53.3 - 60.9) |
| Specificity % (95% CI) | 87.5 (84.2-90.2) | 86.9 (83.5 - 89.7) |
| Indeterminate as negative | ||
| Sensitivity % (95% CI) | 42.7 (38.9-46.6) | 39.3 (35.6 - 43.1) |
| Specificity % (95% CI) | 94.4 (91.9 - 96.3) | 96.8 (94.7 - 98.0) |
Out of the 644 SLE patients, 25 were known to have active lupus nephritis. The clinical sensitivity of the QUANTA Flash dsDNA on this population was 80%, as 20 samples were positive out of the 25. The predicate device detected 20 and 21 samples (with indeterminate results considered as negative or positive, respectively):
| Sensitivity in lupus nephritis: | QUANTA Flash dsDNA | QUANTA Lite dsDNA SC |
|---|---|---|
| Indeterminate as negative | 20/25 (80%) | 20/25 (80%) |
| Indeterminate as positive | 20/25 (80%) | 21/25 (84%) |
Expected values
The expected value in the normal population is "negative". Anti-dsDNA antibody levels were analyzed in a cohort of 300 apparently healthy blood donors (131 females and 169 males, ages 19 to 68 years, with an average and median age of 44 years) using the QUANTA Flash dsDNA. This patient population was different from the one that was used to establish and verify the cutoff. With a cut-off of 27 IU/mL and indeterminate range of 27-35 IU/mL, six samples (2.0%) were in the indeterminate range, and four (1.3%) of the samples were positive with the QUANTA Flash dsDNA. The mean concentration was 11.6 IU/mL, and the values ranged from 666.9 IU/mL). The agreement between QUANTA Flash and the predicate device was 98.8% for the samples below the AMR (4 out of 333 were positive with the QUANTA Lite assay), and it was 100% for samples above the AMR (all samples were positive with the predicate device).
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Additionally, agreement was also calculated on samples that were around the cut-off with the QUANTA Flash dsDNA (25-40 IU/mL). There were 124 samples in this category. The agreement between QUANTA Flash and the predicate device was 69.4% and 55.6 % when indeterminate results were considered as negative and positive, respectively, as shown below:
| Method Comparison -
Within AMR (N=124)
Indeterminate = negative | QUANTA Flash® dsDNA | | | Percent Agreement
(95% Confidence) |
|-----------------------------------------------------------------------|---------------------|-------------|-------|---------------------------------------|
| QUANTA Lite
dsDNA SC
ELISA | Negative | Positive | Total | |
| Negative | 83 | 27 (21 SLE) | 110 | Pos. Agrmnt = 21.4% (7.6 – 47.6%) |
| Positive | 11 (8 SLE) | 3 | 14 | Neg. Agrmnt = 75.5% (66.6 – 82.5%) |
| Total | 94 | 30 | 124 | Total Agrmnt = 69.4% (60.8 – 76.8%) |
| Method Comparison - | QUANTA Flash® dsDNA | Percent Agreement | |||
|---|---|---|---|---|---|
| Within AMR (N=124) | Negative | Positive | Total | (95% Confidence) | |
| Indeterminate = positive | |||||
| QUANTA Lite | |||||
| dsDNA SC | |||||
| ELISA | Negative | 15 | 49 (24 SLE) | 64 | Pos. Agreement = 90.0% (79.9 - 95.3%) |
| Positive | 6 (5 SLE) | 54 | 60 | Neg. Agreement = 23.4% (14.7 - 35.1%) | |
| Total | 21 | 103 | 124 | Total Agreement = 55.6% (46.9 - 64.1%) |
The results show that the majority of the samples that were positive with QUANTA Flash but negative with the predicate device were samples from SLE patients.