SARS-CoV-2 IgG/IgM Rapid Test for the diagnosis of COVID-19

SARS-CoV-2 IgG/IgM rapid test is a lateral flow immunoassay intended for the qualitative detection of IgM and IgG antibodies to SARS-CoV-2 in the blood (serum, plasma) from patients suspected of COVID-19. The SARS-CoV-2 IgG/IgM rapid test is an aid in the diagnosis of patients with suspected SARS-CoV-2 infection in conjunction with clinical presentation and the results of other laboratory tests.

Antibody detection tests are hard to develop but easy to administer. Having a reliable home testing kits for people who think they have had COVID-19 will be a game-changer.

Uses of SARS-CoV-2 IgG/IgM Antibody testing

SARS-CoV-2 IgG/IgM antibody testing is used for rapid screening of SARS‐CoV‐2 carriers, symptomatic or asymptomatic, in hospitals, clinics, and test laboratories. In addition to aiding in the diagnosis, it also reveals evidence of the previous infection and is thus useful; 

  1. To know how many asymptomatic cases have occurred in a population
  2. To know the immune status of high-risk individuals including healthcare professionals
  3. To check SARS-Cov-2 vaccines are working as intended during clinical trials
SARS-CoV-2 IgG/IgM assay
SARS-CoV-2 IgG/IgM assay

Results from the SARS-CoV-2 IgG/IgM Rapid Test should not be used as the sole basis for diagnosis.

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. Positive results for both IgG and IgM could occur after infection and can be indicative of acute or recent infection.

SARS-CoV-2 IgG and IgM in Blood
Variation of the Levels of SARS-CoV-2 RNA and Antigen, IgM and IgG after infection.
(Source: diazyme)

Everybody, especially medical personnel want to know if they already had COVID-19, and is therefore probably immune to infection and can continue their work without fear of infection.

Sensitivity and Specificity of SARS-CoV-2 IgG/IgM assay

The evidence regarding SARS-CoV-2 IgG/IgM assay is currently scarce. Zhentgu et. al., reported overall testing sensitivity of 88.66% and specificity of 90.63% for IgG/IgM assay. This means out of 100 people having COVID-19, this test will show positive results only for 89 people (i.e., 11 people will have false-negative results). Similarly, if we test 100 COVID-19 negative people with this test system, it will give positive results for approximately 9 people (i.e., 9 people will have false-positive test status).

We need more research and publication to establish the diagnostic accuracy of SARS-CoV-2 IgG/IgM assay.

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 SARS-CoV-2 IgG/IgM rapid test early after the infection is unknown.

There are many proteins (antigens) of SARS-CoV-2 and each elicit antibody responses to some extent. So far, we have little knowledge regarding which antigens are best on mounting immune response. Currently, spike protein and nucleocapsid protein are chosen as a suitable candidate. If the chosen protein antigen is not unique, false-positive results will come from people who are immune to other coronaviruses, for example, SARS.

False-positive results for IgM and IgG antibodies may occur due to cross-reactivity from pre-existing antibodies (e.g. previous infection with other coronaviruses which have similar antigenic structure) or other possible causes. At this time, it is unknown for how long IgM or IgG antibodies may persist following infection.

Some scientist argues that once people produce antibodies against a particular coronavirus, they probably have immunity for life.

Materials

The test kit comes with a test cartridge, sample dilution buffer, and a package insert. It also contains external positive and negative controls, to monitor for failures of antibody detection reagents and reaction conditions:

  • Positive Control: Spiked, chemically inactivated, human serum containing IgM and IgG antibodies against SARS-CoV-2 close to the cutoff of the test. Both test lines and the control line should appear.
  • Negative Control: Previously characterized, chemically inactivated, negative human serum. Only the control line should appear.

Cassette design for the detection of COVID-19 Antibodies

A typical immunochromatographic test strip for the detection of IgG or IgM antibodies formed against SARS-CoV-2 consists of;

  1. Sample well: an absorbent pad onto which specimen is applied,
  2. Conjugate or reagent pad that contains SARS-CoV-2 recombinant antigens conjugated with colloidal gold (SARS-CoV-2 conjugates) and rabbit IgG-gold conjugates
  3. Nitrocellulose membrane strip containing the test line and control line.
    1. Test line:
      1. IgG line: immobilized anti-human IgG
      2. IgM line: immobilized anti-human IgM
    2. Control line: immobilized goat anti-rabbit IgG
  4. Waste reservoir composed of another absorbent pad (wick assembly) designed to draw the sample across the reaction membrane by capillary action.

Principle

SARS-CoV-2 IgM/IgG Assay

SARS-CoV-2-IgG/IgM assay is a lateral flow immunoassay used in diagnostic laboratories. For the detection of IgG and IgM antibody against SARS-CoV-2.

When the test specimen is dispensed into the sample well of the test cassette, the specimen migrates by capillary action along the cassette. When the fluid reaches a conjugate pad, the test antibody if present in the specimen, antigen-antibody-conjugate complex is formed.

  1. Anti-SARS-CoV-2 IgG, if present in the specimen: Formation of SARS-CoV-2 Conjugate-IgG complex (i.e., Human IgG-SARS-CoV-2-Ag-SARS-CoV-2-Ab (rabbit)-colloidal gold)
  2. Anti-SARS-CoV-2 vIgM, if present in the specimen: Formation of SARS-CoV-2 Conjugate-IgM complex (i.e., Human IgM-SARS-CoV-2-Ag-SARS-CoV-2-Ab (rabbit)-colloidal gold)

SARS-CoV-Ab-conjugate complex continues to migrate across the membrane until it reaches the capture zone where the complex will bind to immobilized antibodies.

  1. IgG test line: This test line contains anti-human-IgG, so it will only bind with SARS-CoV-2-conjugate-IgG complex.
  2. IgM test line: This test line contains anti-human-IgM, so it will only bind with SARS-CoV-2-conjugate-IgM complex.
Schematic diagram for SARS-CoV-2 IgG/IgM assay
Schematic diagram for SARS-CoV-2 IgG/IgM assay

As more and more Ag-Ab complexes are captured at the “test” lines, the line becomes visible (red line) on the membrane depending on the presence and amount of antibodies in human blood. If the specimen does not contain SARS‐CoV‐2 antibodies, no labeled complexes bind at the test zone and no lines could be observed.

The sample then migrates further along the strip until it reaches the control zone where excess conjugate binds with anti-rabbit IgG (remember; conjugate contains SARS-CoV-2 rabbit IgG, and control line contains anti-goat-anti-rabbit-IgG) producing visible line (control line) on the membrane. This control line indicates that the sample has migrated across the membrane as intended. The presence of a “control” line only indicates that the test was performed properly.

Procedure

  • Allow the device cassette, specimen, and buffer solution to equilibrate to room temperature.
  • Transfer 10 μl of the specimen to the center of the sample well.
  • After the sample well is free of liquid, transfer two drops of simple diluent into the sample well.
  • Wait for 15-20 minutes and read the results.
SARS-CoV-2-IgG/IgM assay procedure and interpretation
SARS-CoV-2-IgG/IgM assay procedure and interpretation
(Image source: Cellex)


Interpretation of COVID-19 serological (IgG/IgM) test

Colored band on IgM test lineColored band on IgG test lineColored band on Control lineResultInterpretation
YesYesYesBoth IgM and IgM against SARS-CoV-2 presentInfection with SARS-CoV-2. Presence of IgM indicates its recent one, and presence of IgG indicates class switching already occurred.
YesNoYesOnly IgM against SARS-CoV-2 present (indicates recent infection)Recent infection with SARS-CoV-2
NoYesYesOnly IgG against SARS-Cov-2 presentPrior infection with SARS-CoV-2 (person is immune to SARS-CoV-2)
Yes/NoYes/NoNoTest invalidIrrespective of presence of test lines, if colored band does not appear in control line, test should be regarded as invalid.

Information regarding the immune response to SARS-CoV- 2 is limited and still evolving. At this time, it is unknown how long IgM or IgG antibodies may persist following infection.

References and further readings

  1. COVID-19, lgG/lgM Rapid Test, Antibody Detection, Blood Test, Serological Test, Coronavirus Disease, Lateral Flow Immunoassay. (n.d.). Retrieved April 14, 2020, from https://cellexcovid.com/
  2. Li, Z., Yi, Y., Luo, X., Xiong, N., Liu, Y., Li, S., Sun, R., Wang, Y., Hu, B., Chen, W., Zhang, Y., Wang, J., Huang, B., Lin, Y., Yang, J., Cai, W., Wang, X., Cheng, J., Chen, Z., … Ye, F. (n.d.). Development and clinical application of a rapid IgM-IgG combined antibody test for SARS-CoV-2 infection diagnosis. Journal of Medical Virology, n/a(n/a). https://doi.org/10.1002/jmv.2572
  3. Why Do We Need Antibody Tests for COVID-19 and How to Interpret Test Results. (n.d.). Retrieved April 14, 2020, from http://www.diazyme.com/covid-19-antibody-tests
  4. Petherick, A. (2020). Developing antibody tests for SARS-CoV-2. The Lancet, 395(10230), 1101–1102. https://doi.org/10.1016/S0140-6736(20)30788-1
  5. Zhao, J., Yuan, Q., Wang, H., Liu, W., Liao, X., Su, Y., Wang, X., Yuan, J., Li, T., Li, J., Qian, S., Hong, C., Wang, F., Liu, Y., Wang, Z., He, Q., Li, Z., He, B., Zhang, T., … Zhang, Z. (2020). Antibody responses to SARS-CoV-2 in patients of novel coronavirus disease 2019. Clinical Infectious Diseases: An Official Publication of the Infectious Diseases Society of America. https://doi.org/10.1093/cid/ciaa344
About Acharya Tankeshwar 445 Articles
Hello, thank you for visiting my blog. I am Tankeshwar Acharya. Blogging is my passion. I am working as an Asst. Professor and Microbiologist at Department of Microbiology and Immunology, Patan Academy of Health Sciences, Nepal. If you want me to write about any posts that you found confusing/difficult, please mention in the comments below.

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