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SARS-CoV-2 Serology (COVID-19) Antibody (IgG/IgM), Immunoassays

Test code(s) 39504, 31672

Question 1. What types of immunoassays are available for SARS-CoV-2 (COVID-19) antibody testing?  

Currently, 2 types of serologic assays are available in the marketplace for SARS-CoV-2 antibody testing:

  1. Laboratory-based immunoassays: Several types of immunoassays are available in this category, such as enzyme-linked immunosorbent assays (ELISAs), chemiluminescent microparticle immunoassays (CMIAs), and immunometric assays.1,2 These tests can be qualitative, semi-quantitative, or quantitative and are generally performed using serum.2  These immunoassays use a solid phase coated with viral antigen to bind SARS-CoV-2 antibodies, but different tests may use different solid phases. For example, ELISAs use a plate and CMIAs use paramagnetic microparticles.1 Quest Diagnostics currently offers high-complexity laboratory-based qualitative immunoglobulin G (IgG) and a duplex IgG/Immunoglobulin M (IgM) SARS-CoV-2 immunoassay. Our IgG and combination IgG/IgM testing are amenable to high-throughput methodologies commensurate with national pandemic testing needs.
  2. Rapid diagnostic tests (RDT)s: These are typically lateral flow assays that can be used for point-of-care (POC) testing. RDTs most frequently test for SARS-CoV-2 IgG, IgM.1 These tests usually use blood samples from a finger stick, and some can use saliva or other specimen types.1,2

Question 2. What is the US Food and Drug Administration (FDA) policy for commercial manufacturers and laboratory development/use of serology tests for SARS-CoV-2?

The FDA’s first guidance in early spring of 2020 to the developers of SARS-CoV-2 antibody tests was more flexible than for molecular tests. On May 4, 2020, the FDA revised this policy to require antibody test developers to submit requests for emergency use authorization (EUA), with their validation data, within 10 business days from the date they notified the FDA of their validation testing OR from the date of this policy, whichever is later.2 The FDA provided specific performance threshold recommendations for specificity and sensitivity.2

SARS-CoV-2 IgG and IgM antibody assays offered by Quest have been authorized through an EUA process.3-6 The FDA has created a website to summarize the performance of the serology tests that have been granted EUA.7

The FDA also issued a letter for healthcare providers on the use of serologic (antibody) tests for COVID-19 on April 17, recommending continued use of serologic tests, as appropriate, with awareness of their limitations.8 Refer to Question 10 for more information about cross-reactivity due to past or present infection with other coronaviruses.

Question 3. How does Quest ensure that the serologic assays are accurate?

Before patient sample testing, Quest verifies the performance characteristics of the FDA EUA designated assays by completing CLIA/CAP-specified in-laboratory verification using stringent acceptability criteria for precision, reproducibility, accuracy, method comparison, cross-reactivity, and clinical performance. Highlights of the manufacturers’ clinical performance of the SARS-CoV-2 IgG and IgM kits used at Quest are as follows:  

  1. SARS-CoV-2 IgG:
    1. Estimated assay sensitivity is 90% to 100% for samples collected at least 15 days post-symptom onset,3-5 based on positive percent agreement (PPA)7 of SARS-CoV-2 IgG serology results on COVID-19 RNA positive patients.3-5
    2. Estimated assay specificity is >99%,3-5 based on negative percent agreement (NPA)7 assessed by performing cross-reactivity studies utilizing serum samples positive for antibodies to other respiratory viruses pre- and post-COVID-19 time periods.3-5
  2. SARS-CoV-2 IgM:
    1. Estimated assay sensitivity is 95% for samples collected at least 15 days post-symptom onset,6 based on PPA7 of SARS-CoV-2 IgM serology results on SARS-CoV-2 RNA positive patients.6
    2. Estimated specificity is >99%6 based on NPA7  assessed by performing SARS-CoV-2 IgM tests on serum samples collected from patients during a pre-COVID-19 time period.6

Question 4. How soon do IgM and IgG antibodies to SARS-CoV-2 appear in a patient who has been infected with the virus?

The antibody response to SARS-CoV-2 usually starts with IgM and/or IgA being detectable first, followed by the longer-lasting and more specific IgG. Data suggest that IgM antibodies can be detected within a few days and IgG antibodies will be detectable from 10 days after COVID-19 symptom onset.3-5 after SARS-CoV-2 exposure or symptom onset.9,10 However, some people do not generate detectable IgG antibodies after infection, because of an underlying immune disorder, immunosuppression, or other, as yet unidentified, reasons.9,10 Additionally, an individual’s immune response can vary in the speed and strength of IgM and IgG production upon exposure to SARS-CoV-2, based on infective dose, viral burden, or other host factors.9,10

Question 5. Can antibody tests be used to diagnose SARS-CoV-2 infection?

No. At present, no antibody tests have an intended use that includes definitive diagnosis of, or ruling out of, current SARS-CoV-2 infection.2,3-6 Serologic IgM and IgG assays provide information about whether a person has been infected with SARS-CoV-2.2,3-6 A molecular diagnostic RNA assay should be considered to diagnose or rule out current infection.2 Although the FDA has not authorized the use of antibody tests to diagnose SARS-CoV-2 infection, serologic assays may be used to support clinical assessment of:

  • Persons who present late in their illnesses (9–14 days after illness onset), when used in conjunction with viral detection tests11; and
  • Persons suspected to have a post-infectious syndrome caused by SARS-CoV-2 infection (eg, multisystem inflammatory syndrome in children [MIS-C]).11

Question 6. How do I interpret SARS-CoV-2 Serology (COVID-19) Antibody (IgG), Immunoassay results?

Results from this qualitative test for SARS-CoV-2 IgG can be positive (reactive), negative (non-reactive), or, occasionally, equivocal (borderline).12-14

A positive (reactive) result indicates detection of SARS-CoV-2 IgG, which may suggest infection with SARS-CoV-2. It usually takes at least 10 days after symptom onset for IgG to reach detectable levels. Although the relationship between IgG positivity and immunity to SARS-CoV-2 has not yet been established, detection of IgG may suggest an immune response to SARS-CoV-2 after resolution of infection. It is currently not known for how long IgG remains detectable after exposure to SARS-CoV-2.12-14

A negative (non-reactive) result indicates that SARS-CoV-2 IgG is not present at a level that is detectable by the SARS-CoV-2 Serology (COVID-19) Antibody (IgG), Immunoassay.12-14 Negative results suggest that a person has not been infected with SARS-CoV-2 or has been very recently infected (antibodies have not yet been produced).12-14 Such a patient is at risk of infection, however, the rate of IgG development varies between individuals. A detectable IgG response usually takes at least 10 days after exposure but may take up to 3 weeks or longer to develop in some patients, and some people may not develop antibodies at all. The IgG response is usually slower in people who are either taking immunosuppressive therapies or are immunocompromised because of a health condition.12-14 Given the variability in timing of an IgG response, a repeat serology IgG test should be considered after an additional 2- to -3-week period for patients who are initially antibody negative and are thought to have clinically recovered from a SARS-CoV-2 infection.12-14

An equivocal (borderline) result indicates that IgG was detected at a level close to the threshold of the limit of detection for the test.12-14 Equivocal results may occur for unidentified reasons or may represent an early infection, detection of an IgG generated from a past infection (refer to Question 9 for cross-reactivity with non-SARS-CoV-2 coronavirus strains), or, in some cases, an underlying immune disorder, immunosuppression, or other conditions.12-14

In populations with a high prevalence of infection with non-SARS-CoV-2 coronavirus strains, a positive result could also be due to past or present infection with one of these strains (refer to Question 10).

Question 7. How do I interpret SARS-CoV-2 Serology (COVID-19) Antibody (IgM), Immunoassay results?

Results from this qualitative test for SARS-CoV-2 IgM can be positive (reactive) or negative (non-reactive).15

A positive (reactive) result indicates that antibodies to SARS-CoV-2 were detected and the individual has potentially been infected with SARS-CoV-2.15 SARS-CoV-2 IgM is generally detectable in blood several days after initial infection. Detection of IgM may indicate a recent infection.15 It is unknown how long SARS-CoV-2 IgM remains detectable post-infection.11,15

Whether SARS-CoV-2 antibodies confer immunity to infection also remains unknown.11,15 Incorrect assumptions of immunity may lead to premature discontinuation of physical distancing requirements and could increase the risk of infection for individuals, their household members, and the public.15

A negative test result means that SARS-CoV-2 IgM was not present in the specimen at levels above the limit of detection.6 However, patients tested early after infection may not have detectable antibodies despite active infection, and patients tested X days after infection will not have detectable IgM. A negative result should not be used to rule out infection. Testing of patient samples for the presence of virus material (eg, RNA or antigen) should be performed if acute infection is suspected.15

Regardless of the test result, individuals should continue to follow CDC guidelines to reduce the risk of infection, including social distancing and wearing masks.15

Question 8. Does a positive result for IgG and/or IgM antibody to SARS-CoV-2 mean that the patient is immune to reinfection? 

The presence of IgG or IgM antibody to SARS-CoV-2 indicates that the patient has mounted an immune response to the virus. Although the immune response may protect against reinfection, this has yet to be conclusively established.12 It is not known how long antibodies to the virus will protect someone, if at all.12  Cases of reinfection with COVID-19 have been reported but remain rare​.12​ Based on what we know from similar viruses, some reinfections are expected. Scientists are conducting research to answer these questions.12

Question 9. What are the indications for use of the SARS-CoV-2 Serology (COVID-19) Antibody (IgG/IgM), Immunoassay?

Testing for SARS-CoV-2 IgG/IgM, including for individuals who may be asymptomatic or are ≥10 days after SARS-CoV-2 exposure or post-symptom onset, can play an important role in the fight against COVID-19.13 SARS-CoV-2 IgG/IgM testing can be used to

  • Identify SARS-CoV-2-infected persons with NAAT/PCR-negative results, especially patients who present late with RNA levels below the detection limit of a NAAT/PCR assays, or when respiratory tract sampling is not possible.2,7-10,13
  • Identify whether people have been infected with SARS-CoV-2 and have mounted an immune response.2,7,8,11,13
  • Assess how many people have been infected with SARS-CoV-2 in a population, by identifying individuals who have developed antibodies to the virus.2,7,8,11,13
  • Possibly help identify individuals who may be able to donate convalescent plasma as a possible treatment for those who are seriously ill from COVID-19.2,7,8,11,13

In the future, this test, along with other clinical data, may help identify individuals who may be less susceptible to reinfection.

Question 10. What proportion of the general population has pre-existing IgM and/or IgG reactivity to SARS-CoV-2 antibody tests?

Manufacturers of the SARS-CoV-2 IgG test performed by Quest using validation samples from pre-COVID times (2010, 2017, and 2019), had less than 1% of samples showing positive results.3-5

In an antibody cross-reactivity study of 143 specimens from 30 different categories of infectious disease and autoimmune disease, only 1 sample with rheumatoid factor reported a SARS-CoV-2 IgM positive result.6 In a medical condition cross-reactivity study using 65 specimens from 13 different categories, only 1 sample from a hemodialysis patient tested positive for SARS-CoV-2 IgM.6

The cross-reactivity for Quest SARS-CoV-2 IgG (ELISA or CMIA) and SARS-CoV-2 IgM Immunoassays (CMIA) is <%.3-6

Question 11. Can I simultaneously test for SARS-CoV-2 IgM and IgG serology at Quest?

Yes. Test code 31672 qualitatively assesses and reports the presence or absence of SARS-CoV-2 IgM and IgG antibodies in patient serum specimens.

Question 12. What is the utility of simultaneously testing for SARS-CoV-2 IgM and IgG? 

In a review of current advances in SARS-CoV-2 serology testing (32 peer-reviewed and 23 preprint publications), the combined use of IgM and IgG detection resulted in a higher sensitivity than that observed when detecting either antibody isotype alone.9 The same study concluded that simultaneous IgG/IgM measurements can improve sensitivity of SARS-CoV-2 RNA testing protocols in early post-onset of symptoms.9

Question 13. What is the role of antibody neutralization assays, and are they available to assess immunity to SARS-CoV-2?

Neutralization assays are done to help assess the ability of neutralizing antibodies (NAbs) in patient serum to neutralize virus infectivity.16 These tests are useful to assess whether past infection (or vaccination, if available) has provided protection against infection. In these tests, serial dilutions of patient serum are incubated with known infectious virus concentrations and then added to virus susceptible cell lines.16 After incubation, infected cells are quantified to determine the effectiveness of patient antibodies in neutralizing the virus.16,17 The presence of NAbs may suggest in vivo immunity to viral infection, and convalescent serum with NAbs is being studied as a treatment for severe COVID-19.16-19 Testing for NAbs is only available at some specially equipped public health laboratories and research facilities. It is not performed at Quest.

Question 14. Are SARS-CoV-2 qualitative IgM and/or IgG tests helpful in assessing immunity after vaccination?

At the date of this publication, no qualitative SARS-CoV-2 IgG and IgM assays, under FDA EUA, have an intended use for evaluating SARS-CoV-2 vaccine efficacy.

Clinical researchers continue to investigate what constitutes SARS-CoV-2 protective immunity and vaccine efficacy.

References

  1. Kobokovich A, West R, Gronvall G. Serology based tests for COVID-19. Center for Health Security. Accessed October. 25, 2020. https://www.centerforhealthsecurity.org/resources/COVID-19/serology/Serology-based-tests-for-COVID-19.html
  2. FAQs on testing for SARS CoV-2. US Food and Drug Administration. Accessed October 25, 2020. https://www.fda.gov/medical-devices/emergency-situations-medical-devices/faqs-diagnostic-testing-sars-cov-2
  3. Anti-SARS-CoV-2 ELISA (IgG). Instructions for use. Euroimmun; 2020.
  4. Abbott Architect SARS-CoV-2 IgG. Package insert. Abbott Laboratories; 2020.
  5. VITROS Immunodiagnostic Products Anti-SARS-CoV-2 IgG. Instructions for use. Ortho-Clinical Diagnostics; 2020.
  6. Abbott AdviseDx SARS-CoV-2 IgM. Package insert. Abbott Laboratories; 2020. Accessed October 23, 2020. https://www.fda.gov/media/142940/download
  7. EUA authorized serology test performance. US Food and Drug Administration. Updated October, 14, 2020. Accessed October 23, 2020. https://www.fda.gov/medical-devices/emergency-situations-medical-devices/eua-authorized-serology-test-performance
  8. Important information on the use of serologic (antibody) tests for COVID-19–letter to health care providers. US Food and Drug Administration. Updated April 17, 2020. Accessed October 25, 2020. https://www.fda.gov/medical-devices/letters-health-care-providers/important-information-use-serological-antibody-tests-covid-19-letter-health-care-providers
  9. Espejo AP, Akgun Y, Al Mana AF, et al. Review of current advances in serologic testing for COVID-19. Am J Clin Pathol. 2020;154(3):293-304. doi:10.1093/ajcp/aqaa112
  10. Li Z, Yi Y, Luo X, et al. Development and clinical application of a rapid IgM‐IgG combined antibody test for SARS‐CoV‐2 infection diagnosis. J Med Virol.2020;10.1002/jmv.25727. doi:10.1002/jmv.25727
  11. Interim guidelines for COVID-19 antibody testing. Centers for Disease Control and Prevention. Updated August 1, 2020. Accessed October 25, 2020.https://www.cdc.gov/coronavirus/2019-ncov/lab/resources/antibody-tests-guidelines.html#anchor_1590264293982
  12. Fact sheet for healthcare providers: SARS-Co-V-2 IgG assay - Abbott Laboratories Inc. US Food and Drug Administration. Updated April 26, 2020. Accessed November 5, 2020. https://www.fda.gov/media/137381/download
  13. Fact sheet for healthcare providers: EUROIMMUN Anti-SARS-CoV-2 ELISA (IgG). US Food and Drug Administration. Updated May 4, 2020. Accessed November 5, 2020. https://www.fda.gov/media/137607/download
  14. Fact sheet for healthcare providers: Ortho-Clinical Diagnostics, Inc. VITROS Immunodiagnostic Products Anti-SARS-CoV-2 IgG Reagent Pack. US Food and Drug Administration. Updated October 23, 2020. Accessed November 5, 2020. https://www.fda.gov/media/137361/download
  15. Fact Sheet for Healthcare Providers: Abbott Laboratories AdviseDx SARS-CoV-2 IgM. US Food and Drug Administration. Updated October 9, 2020. Accessed October 26, 2020. https://www.fda.gov/media/142938/download
  16. Reinfection with COVID-19. Centers for Disease Control and Prevention. Updated October 27, 2020. Accessed November 5, 2020. https://www.cdc.gov/coronavirus/2019-ncov/your-health/reinfection.html
  17. Ju B, Zhang Q, Ge J, et al. Human neutralizing antibodies elicited by SARS-CoV-2 infection. Nature. 2020;584(7819):115-119. doi:10.1038/s41586-020-2380-z
  18. Mo H, Zeng G, Ren X, et al. Longitudinal profile of antibodies against SARS-coronavirus in SARS patients and their clinical significance. Respirology. 2006;11(1):49–53. doi:10.1111/j.1440-1843.2006.00783.x
  19. IDSA COVID-19 antibody testing primer. Infectious Diseases Society of America. Updated April 22, 2020. Accessed April 23, 2020. https://www.idsociety.org/globalassets/idsa/public-health/covid-19/idsa-covid-19-antibody-testing-primer.pdf
This FAQ is provided for informational purposes only and is not intended as medical advice. A clinician’s test selection and interpretation, diagnosis, and patient management decisions should be based on his/her education, clinical expertise, and assessment of the patient.
Document FAQS.219 Version: 2
Version 2: Effective 11/9/2020 to present
Version 1: Effective 05/26/2020 to 11/9/2020
Version 0: Effective 04/24/2020 to 05/26/2020