Artron COVID-19 IgM/IgG Antibody Test Instruction Manual

Artron COVID-19 IgM/IgG Antibody Test

INTENDED USE

Artron COVID-19 IgM/IgG Antibody Test is a rapid and qualitative immunochromatographic in-vitro assay for the differential detection of IgM &/or IgG antibodies to SARS-CoV-2 in human serum, plasma, venous whole blood or fingerstick whole blood samples obtained from patients with suspected SARS- CoV-2 infection. The device is designed to aid in the determination of recent or previous exposure to the SARS-CoV-2 virus, assessing the development of an immune response to the virus after SARS-CoV-2 infection. The test may also detect a response to vaccination but is unable to distinguish between previously infected individuals and vaccinated individuals. This assay is not intended to be used for screening patients or as an aid for the diagnosis of patients with suspected COVID-19 infection. This assay is not intended for home testing (or self-testing). A positive result does not necessarily mean a current infection but represents a different stage of the disease after infection. Current infection cannot be established by this device and must be confirmed by Real-Time Reverse Transcriptase Polymerase Chain Reaction (RT- PCR) or viral gene sequencing. Negative results do not preclude SARS-CoV-2 infection and should not be used as the sole basis for patient management decisions. Negative results must be combined with clinical observations, patient history, and epidemiological information. False-negative results can occur in elderly and immunocompromised patients. False positive results from IgM and IgG antibodies may occur due to cross-reactivity from pre-existing antibodies or other possible causes. The test is intended for professional use or point-of-care use and should only be used by healthcare professionals trained in the techniques of in vitro diagnostics procedures and proper infection control procedures.

SUMMARY AND PRINCIPLE OF THE ASSAY

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the virus strain that caused an outbreak of a novel coronavirus disease (COVID-19), which has subsequently affected countries and regions worldwide. Severe disease onset might result in death due to massive alveolar damage and progressive respiratory failure. On March 11, 2020, the World Health Organization (WHO) has declared the global outbreak of COVID-19 a pandemic associated with substantial morbidity and mortality. The principle of Artron COVID-19 IgM/IgG Antibody Test is an antibody-capture immunochromatographic assay for the simultaneous detection and differentiation of IgM & IgG antibodies to SARS-CoV-2 virus in human serum, plasma, or whole blood samples. SARS-CoV-2-specific antigens are conjugated to colloidal gold and deposited on the conjugate pad. Monoclonal anti-human IgM and monoclonal anti-human IgG are immobilized on two individual test lines (M line and G line) of the nitrocellulose membrane. The M line is closer to the sample well and is followed by the G line. When the sample is loaded, the gold-antigen conjugate is rehydrated and the SARS-CoV-2IgM and/or IgG antibodies, if any in the sample, will interact with the gold-conjugated antigen. The immunocomplex will migrate towards the test window reaching the test zone (G line & M line) where they will be captured by the relevant anti-human IgM (M line) and/or anti- human IgG (G line), forming a visible pink line, indicating positive results. If SARS-CoV-2 antibodies are absent in the sample, no pink line will appear in the test lines (G line & M line), indicating a negative result. To serve as internal process control, a control line should always appear at Control Zone (C line) after the test is completed. The absence of a pink control line in the Control Zone is an indication of an invalid result.

PACKAGE CONTENTS

• Pouch contents: One test cassette with test strip, One desiccant;
• 25 capillary tubes for 25 tests (to collect fingerstick blood);
• Sample buffer 3 ml/vial for 25 tests.

MATERIALS REQUIRED (BUT NOT PROVIDED)

• Alcohol swab
• Safety lancet (for fingerstick whole blood specimens)
• Blood collection device. (for other than fingerstick whole blood specimens)
• Precision pipette capable of delivering 10μl and/or 20μl with disposable tips (for other than fingerstick whole blood specimens)
• Gloves
• Clock or timer

WARNINGS AND PRECAUTIONS

• For professional in-vitro diagnostic use only.
• Do not reuse.
• Do not use if the product seal or its packaging is compromised.
• Do not use after the expiration date shown on the pouch.
• Do not mix and interchange different specimens.
• This test should be performed at 2 to 30°C (17 to 86°F). If stored refrigerated, ensure that the Test Units are brought to operating temperature before performing testing.
• Wear protective clothing such as laboratory coats, disposable gloves and eye protection while handling potentially infectious materials or performing the assay.
• Wash hands thoroughly after finishing the tests.
• Do not eat, drink, or smoke in the area where the specimens or test are handled.
• Clean up spills thoroughly with appropriate disinfectants.
• Handle all specimens as if they contain infectious agents. Observe established precautions against microbiological hazards throughout testing procedures.
• Dispose of all specimens and used devices in a proper bio-hazard container. The handling and disposal of hazardous materials should follow local, regional, or national regulations.
• Keep out of children’s reach.
• For additional information on COVID-19 antibody (serology) testing: Information for patients, please refer to Health Canada for COVID-19 antibody (serology) testing:
• Information for patients: https://www.canada.ca/en/health-canada/services/drugs-health-products/covid19-industry/medical-devices/testing/serological/information-for-patients.html

SPECIMEN PREPARATION

• Whole Blood samples may be collected by fingerstick or venipuncture, following routine facility procedures. In summary:

Fingerstick whole blood

• Clean the area of finger to be lanced with the alcohol swab. Allow to dry.
• Without touching the puncture site, rub down the hand towards the middle or ring finger fingerstick.
• Puncture the skin with a sterile lancet and wipe away the first drop of blood.
• Gently rub the hand from wrist to the lanced finger to form a full drop of blood over the puncture site.
• Collect the blood droplet using the included capillary tube.
• Fingerstick whole blood must be tested immediately after collection.

Venous whole blood

• Collect venous whole blood in a tube with an anticoagulant (Heparin/EDTA/CPDA-1).
• Serum samples, collect blood in a tube without anticoagulant and allow it to clot.
• Plasma samples, collect blood in a tube containing an anticoagulant (Heparin/EDTA/CPDA-1).
• Separate serum or plasma from blood as soon as possible to avoid hemolysis. Use only clear, non-hemolyzed specimens.
• Whole blood in blood collection tubes with anticoagulant (Heparin/EDTA/CPDA-1), serum/plasma in blood collection tubes may be stored at 2ºC to 8ºC for up to 7 days, if the sample needs to be transported or the tests cannot be performed immediately. Bring the sample to attain room temperature (without heating) prior to use.

TEST PROCEDURES

1. Get the test cassette from the sealed pouch by tearing at the notch and place the cassette on a flat, dry surface.
2. For fingerstick or venous whole blood: Using a capillary tube, collect the fingerstick whole blood or venous whole blood till the black line. (20μl) For serum/plasma: Using a pipette, collect the serum/plasma (10μl).
3. Add the collected serum/plasma/whole blood to the upper area (close to test window) of the sample well on the test cassette without air bubbles (hold the capillary tube/ pipette vertically and gently touch the end against the pad within the sample well for transferring).
4. Wait for 20-30 seconds; add 2 drops (around 90μl) of sample buffer to the sample well of the test cassette.
5. Wait and read the results in 15-30 minutes.

WARNING: DO NOT INTERPRET RESULTS AFTER 30 MINUTES.

RESULT INTERPRETATION

Negative

• A pink-colored band appears only at the control region (C), indicating a negative result for SARS-CoV-2 infection.

Positive

Pink-colored bands appear at the control region (C) and G and /or M region.

1. IgM and IgG Positive, visible bands at both M and G.
2. IgM positive, a visible band at M region.
3. IgG positive, a visible band at G region.

Invalid

• No visible band at the control region (C). Repeat with a new test. If test still fails, please contact the distributor with the lot number.

QUALITY CONTROL

Although the testing cassette contains an internal quality control (pink colored band in the control region), good laboratory practice recommends the daily use of an external control to ensure proper testing performance. Quality control samples should be tested according to the standard quality control procedure established by your laboratory.

STORAGE AND STABILITY

• The test should be stored at 2-30ºC in its original package, avoid direct sunlight and freezing.
• Opened test cassette should be used within 4 hours.
• Do not freeze the test device.
• Shelf life: 18 months

LIMITATIONS

• The instructions for use of the test should be followed during testing procedures.
• Use in conjunction with the testing strategy outlined by public health authorities in your area.
• Humidity and temperature can adversely affect the results.
• There is always a possibility that false results will occur due to the presence of interfering substances in the specimen or factors beyond the control of the manufacturer, such as technical or procedural errors associated with the testing.
• The reagent can only be used to determine the immune status of the body to SARS-CoV-2 after infection, but not directly to diagnose current SARS-CoV-2 infection.
• Negative results do not preclude SARS-CoV-2 infection and should not be used as the sole basis for patient management decisions. IgM antibodies may not be detected in the first few days of infection; the sensitivity of the test early after infection is unknown.
• Results are for the detection of SARS-CoV-2 antibodies. IgM antibodies to SARS-CoV-2 are generally detectable in blood several days after initial infection, although levels over the course of infection are not well characterized. IgG antibodies to SARS-CoV-2 become detectable later following infection. At this time, it is unknown how long IgM or IgG antibodies may persist following infection.
• Positive results for both IgG and IgM could occur after infection and can be indicative of acute or recent infection (and successful immune response to a vaccine, once developed).
• False positive results for IgM and IgG antibodies may occur due to cross-reactivity from pre-existing antibodies or other possible causes.
• False-negative results for IgM and IgG antibodies may occur due to high-dose hook effect. It is recommended to further dilute the specimens which are suspected of carrying high-dose hook effect and combine other test results along with comprehensive consideration of clinical history.
• Although Artron COVID-19 IgM/IgG Antibody Test applies multiple antigens to prevent missed detection. However, it still could not rule out the possibility of in false negative test results due to SARS-CoV-2 variants.
• The presence of specific antibodies is a sign of previous or current infection and can also be used to determine the efficacy of treatment.
• Laboratories are required to report all positive results to the appropriate public health authorities.
• The performance of this device has not been assessed in a population vaccinated against COVID-19.
• The performance of the device has not been assessed on specimens from individuals who have been infected with emerging variants of SARS-CoV-2 of public health concern.
• This test identifies antibodies to the spike protein of the SARS-CoV-2 virus and is therefore unable to distinguish between previously infected individual and vaccinated individuals.

PERFORMANCE CHARACTERISTICS

Analysis of Sensitivity and Specificity

NIBSC Anti-SARS-CoV-2 Verification panel (code: 20/B770) was used to validate the analytic sensitivity of Artron COVID-19 IgM/IgG Antibody Test. Artron COVID-19 IgM/IgG Antibody Test identified all the 23 positive members from 37 panel members including 14/23(60.87%) IgM positive and 23/23 (100%) IgG positive; the analytic sensitivity for IgG of Artron COVID-19 Antibody Test is 12.5U(1:80), the analytic sensitivity for IgM of Artron COVID-19 Antibody Test is 125U (1:8), when testing serially diluted NIBSC diagnostic calibrant working standard (code: 20/162). No false positive results were observed when testing the 14 negative members in the NIBSC Anti-SARS CoV-2 Verification panel (code: 20/B770).

Samples with the below seromarkers associated with unrelated SARS-CoV-2 medical conditions were used to evaluate cross-reaction:

• Rheumatoid factor (RF)
• Anti-nuclear antibodies (ANA)
• Coronavirus Seasonal
• Coronavirus 229E
• Coronavirus NL63
• Coronavirus HKU1
• Coronavirus OC43
• Anti-Human immunodeficiency virus (HIV)
• Human Hepatitis B virus (HBV) serum markers (HBsAg, anti-HBc IgG/IgM)
• Anti-Human Hepatitis C virus (HCV)
• Anti-Helicobacter pylori (HP)
• Anti-Herpes simplex virus (HSV)
• Anti-Cytomegalovirus (CMV)
• Anti-Chikungunya virus (CHIKV)
• Anti-Zika virus (ZIKV)
• Anti-Mycoplasma
• Anti-Dengue virus (DENV)
• Anti-Adenovirus (ADV)
• Anti-Respiratory syncytial virus (RSV)
• Anti-Influenza A
• Anti-Influenza B
• Anti-Parainfluenza virus 1/2/3
• Anti-Epstein-Barr virus (EBV)
• Anti- Human Metapneumovirus (hMPV)
• Anti-Haemophilus influenza
• Anti-Streptococcus pyogenes
• Anti-Mycobacterium tuberculosis (TB)
• Anti-T. pallidum (TP)

Total 143 samples were applied to evaluate the analytical specificity of Artron COVID-19 IgM/IgG Antibody Test. The results showed that only one test gave very equivocal signal on IgM with one anti-CHIKV plasma, and Artron COVID-19 IgM/IgG Antibody Test had no cross reaction with all other 142 samples. No significant cross-reactivity was observed to most of above relevant serum markers. The results demonstrated that COVID-19 IgG/IgM Antibody Test has good analytical specificity.

Interference Study

Analytes commonly found in OTC, prescription and/ or abuse drugs, chemical analytes, and endogenous substances did not interfere with Artron COVID-19 IgM/IgG Antibody Test.

Repeatability and Reproducibility

Tests showed positive results with all positive samples and showed negative results with negative samples. There was no significant difference observed to the same sample when repeatedly testing 10 tests in the same batch. No appreciable intra and inter lot variation were observed among different tests for each lot, different lots, different operators at different test sites in different time for the same sample. The results demonstrated that the repeatability and reproducibility of Artron COVID-19 IgM/IgG Antibody Test is satisfactory.

Diagnostic Sensitivity and Specificity

1. Diagnostic Sensitivity and Specificity of Sera/Plasmas/Venous Whole Blood Specimens

A total of 1863 samples including 686 of molecular diagnosis confirmed SARS- CoV-2 positive sera/plasma/whole blood samples and 1177 of clinically true (collected before Nov. 2019) or RT-PCR confirmed SARS-CoV-2 negative samples sera/plasma samples (not including 19 SARS-COV-1 positive sera) were used to evaluate Artron COVID-19 IgM/IgG Antibody Test from six evaluation centers. Out of all the 686 positive samples, Artron COVID-19 IgM/IgG Antibody Test identified 580 of IgM positive and 570 of IgG positive and 626 of SARS-CoV-2 IgM&/or IgG positive samples. The diagnostic sensitivity for IgM is 84.55%, for IgG is 83.09% and the combined sensitivity is 91.40%. The diagnostic sensitivity of IgG for over 14 days after onset is 94.91%. The diagnostic sensitivity of IgG and IgM combined for over 14 days after onset is 97.22%. The diagnostic specificity for IgM is 98.05%, for IgG is 99.75% and the combined specificity is 97.88%. The overall agreement for IgM and IgG is 93.08% and 93.61%, respectively. The combined overall agreement is 95.44%. Cross-reactivity to SARS-COV-1 was not evident on the IgM portion but there was some reactivity on the IgG portion of the test.

Summary for all the test results of sera/plasma/venous whole blood

Diagnostic Sensitivity of Seroconversion

The seroconversion evaluation sensitivity of the Artron COVID-19 IgG/IgM Antibody Test Cassette was evaluated on 51 serum samples from 8 individuals getting treatment at the Central Laboratory of Saitama Medical University Hospital in Japan. The collected samples were chosen from the patients with the early infection, from one to three weeks after onset. All the studied cases are confirmed by RT-PCR. The anti-SARS-CoV-2 antibodies levels of the patients were examined using Artron COVID-19 IgM/IgG Antibody Test. Artron COVID-19 IgG/IgM Antibody Test detected IgG seroconversion during 9-15 days after onset, in all 8 cases and the seroconversion rate was 100%.

summary for the positive rate in eight SARS-CoV-2 seroconverted patients

After onset

（ day ）

| Artron COVID-19 IgM/IgG Antibody Test
---|---
IgM| IgG
Negative| Positive| Negative| Positive
0-8| 5| 3| 7| 1
9-15| 0| 8| 0| 8
---|---|---|---|---
Positive rate (%)| IgM| IgG
0-8| 37.5| 12.5
9-15| 100| 100

Diagnostic Sensitivity and Specificity of Fingerstick Whole Blood Specimens

A total of 130 samples were collected in point-of-care testing including 70 molecular diagnosis confirmed SARS-CoV-2 positive fingerstick whole blood samples and 60 RT-PCR confirmed SARS-CoV-2 negative samples. Of all the chosen fingerstick whole blood samples, Artron COVID-19 IgM/IgG Antibody Test identified 59 IgM positive and 51 IgG positive cases out of a total of 70 COVID-19 IgM &/or IgG positive from 70 SARS-CoV-2 infected patients’ samples. The diagnostic sensitivity for IgM was 84.29%, for IgG was 72.86%, and the combined sensitivity was 100%; the diagnostic sensitivity for IgG after onset

14 days is 92.16%. Out of a total of 60 negative fingerstick whole blood samples, one IgM false positive case and none IgG false positive case was observed and the diagnostic specificity for IgM was 98.33% and for IgG was 100%; the combined specificity was 98.33%.

BIBLIOGRAPHY

• Clinical management of severe acute respiratory infection (SARI) when COVID-19 disease is suspected. Interim guidance. World Health Organization.13 March 2020.
• Report of the WHO-China Joint Mission on Coronavirus Disease 2019 (COVID-19). World Health Organization. 16-24 February 2020.
• The Epidemiological Characteristics of an Outbreak of 2019 Novel Coronavirus Diseases (COVID-19). Chinese Center for Disease Control and Prevention. CCDC Weekly, 2(8):113- 122, 2020.
• A novel coronavirus outbreak of global health concern. Wang C et al. Lancet, 395(10223): 470- 473, 2020

MANUFACTURER CONTACT INFORMATION

• Artron Laboratories Inc.
• ADDRESS: 3938 North Fraser Way Burnaby, BC V5J 5H6 Canada
• Ph: 604.415.9757
• Fax: 604.415.9795
• www.artronlab.com
• info@artronlab.com

MedNet EC-REP GmbH Borkstrasse 10 48163 Muenster Germany

Doc No. A03-51-322 VER. 08 Revision: Apr. 2021

References

• Artron Laboratories Inc. Official Sit| The Canadian leading IVD manufacture for medical devidce and helath. Government supplier.

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