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SARS-CoV-2 Antibody Testing

Test code(s) 39749, 31672, 34499, 39820

The serology tests offered by Quest for detection of SARS-CoV-2 antibodies are summarized in the Table below:

Quest utilizes only SARS-CoV-2 antibody tests that have received emergency use authorization (EUA) from the US Food and Drug Administration (FDA). At this time, our SARS-CoV-2 antibody tests are laboratory-based, qualitative, and semi-quantitative immunoassays; we do not offer rapid antibody tests.

At present, EUA SARS-CoV-2 antibody tests fall into 2 main groups: 

  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 Within this laboratory-based test group, assays can be qualitative, semi-quantitative, or quantitative and are generally performed using serum.1-3 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 currently offers high-complexity laboratory-based qualitative immunoassays for SARS-CoV-2. One is for immunoglobulin G (IgG) and the other is a duplex assay for SARS-CoV-2 IgG/Immunoglobulin M (IgM). Our IgG and combination IgG/IgM tests are amenable to high-throughput methodologies commensurate with national pandemic testing needs.

  2. Rapid serology/antibody tests: These are typically lateral flow assays that can be used for point-of-care (POC) testing. Rapid antibody tests most frequently test for SARS-CoV-2 IgG and/or IgM.1 These tests usually use blood specimens from a finger stick, and some can use saliva or other specimen types.1

The FDA’s first guidance in early spring of 2020 for 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 EUA requests, 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.3

SARS-CoV-2 IgG, IgM and total antibody assays offered by Quest have been authorized through an EUA process.3-8 The FDA has created a website to summarize the performance of the serology tests that are authorized.On April 17, 2020, the FDA also issued a letter for healthcare providers recommending continued use of serologic tests, as appropriate, with awareness of their limitations.2

Before patient specimen testing, Quest verifies the performance characteristics of the FDA EUA authorized 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 Antibody (IgG), Nucleocapsid, Qualitative ([test code 39749]; also a component of the IgM/IgG panel [test code 31672])
    1.  Estimated assay sensitivity is >99.6% for specimens collected at least 15 days post–symptom onset,4 based on positive percent agreement (PPA)3 of SARS-CoV-2 IgG serology results among SARS-CoV-2 RNA−positive patients.4
    2.  Estimated assay specificity4 is >99.9%, based on negative percent agreement (NPA)3 assessed by performing cross-reactivity studies utilizing serum specimens positive for antibodies to other respiratory viruses pre– and post–COVID-19 time periods.4
  2.  SARS-CoV-2 Serology (COVID-19) Antibodies, IgM (spike), Qualitative (a component of the IgM/IgG panel [test code 31672])
    1. Estimated assay sensitivity is 95% for specimens collected at least 15 days post–symptom onset,5 based on PPA3 of SARS-CoV-2 IgM serology results among SARS-CoV-2 RNA–positive patients.5
    2. Estimated specificity5 is >99%, based on NPA3 assessed by performing SARS-CoV-2 IgM tests on serum specimens positive for antibodies to other respiratory viruses pre– and post–COVID-19 time periods.5
  3. SARS-CoV-2 Antibody (IgG), Spike, Semi-Quantitative [test code 34499]
    1. Estimated assay sensitivity is >99.9% for specimens collected at least 15 days post–symptom onset,6,7 based on PPA3 of SARS-CoV-2 IgG serology results among SARS-CoV-2 RNA−positive patients.6,7
    2. Estimated assay specificity is approximately6,7 99.9%, based on NPA4 assessed by performing cross-reactivity studies utilizing serum specimens positive for antibodies to other respiratory viruses pre– and post–COVID-19 time periods.6,7
  4. SARS-CoV-2 Total Antibody, Spike, Semi-Quantitative [test code 39820]
    1. Estimated assay sensitivity is >99.9% for specimens collected at least 15 days post–symptom onset,8 based on PPA3 of SARS-CoV-2 IgG serology results among SARS-CoV-2 RNA−positive patients.8
    2. Estimated assay specificity is approximately8 99.9%, based on NPA3 assessed by performing cross-reactivity studies utilizing serum specimens positive for antibodies to other respiratory viruses pre– and post–COVID-19 time periods.8

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 post-infection and IgG antibodies will be detectable from some individuals by 10 days after COVID-19 symptom onset.3-8 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 host factors.9,10 In a review of current advances in SARS-CoV-2 serology testing, the combined use of IgM and IgG detection resulted in a higher sensitivity than that observed when detecting either antibody isotype alone.9,10

No. At present, no antibody tests have an FDA EUA specified intended use that includes definitive diagnosis of, or ruling out of, current SARS-CoV-2 infection.2-9 Serologic IgM and IgG assays provide information about whether a person has developed an immune response to a COVID-19 infection.1,3-8 A molecular diagnostic RNA assay should be considered to help diagnose a current infection.1,3-8 Although the FDA has not specified the intended use of antibody tests to diagnose SARS-CoV-2 infection, serologic assays may be used to support clinical assessment of

  • Persons who present to clinicians late in their illness (9-14 days after symptom onset), when used in conjunction with viral detection tests.1

  • Persons suspected to have a post-infectious syndrome caused by SARS-CoV-2 infection (eg, multisystem inflammatory syndrome in children [MIS-C]).1

The COVID-19 vaccines launched in the United States can induce an immune response to the spike (S) protein of SARS-CoV-2.1,11-13 A positive SARS-CoV-2 spike IgG antibody result may reflect an immune response to a recent/prior natural SARS-CoV-2 infection.4,6-8 Because the COVID-19 vaccines launched in the United States an immune response to the spike (S) protein of SARS-CoV-2,1,11-13 a positive SARS-CoV-2 spike IgG antibody result could also reflect an immune response to COVID-19 vaccination. However, the clinical significance of an immune response after vaccination is not yet known.1,11-13 At this time, there are no FDA authorized tests for individuals who have received a COVID-19 vaccination, and the performance characteristics of the authorized antibody tests have not been established for those individuals.3 Similarly, the CDC states, “Antibody testing is not currently recommended to assess for immunity to COVID-19 following COVID-19 vaccination or to assess the need for vaccination in an unvaccinated person.”1

A positive/reactive nucleocapsid antibody test result indicates recent or prior infection with the virus and a potential immune response to COVID-19.1,4,14

A negative result for spike or nucleocapsid SARS-CoV-2 IgG means there is no evidence of an immune response to recent/prior infection.1,4,6,7,14-16   

A nucleocapsid SARS-CoV-2 IgG test would not be expected to detect an immune response in individuals who have received COVID-19 spike vaccination (and who had not been infected naturally).

The IgG response is usually slower in people who are either taking immunosuppressive therapies or immunocompromised because of a health condition.4,11Given 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 individuals who are initially antibody negative and are thought to have clinically recovered from a SARS-CoV-2 infection.4,11

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. For more information on common coronavirus surveillance information, refer to the CDC Surveillance for Common Human Coronaviruses webpage: CDC.gov/Surveillance/REVSS/Coronavirus/Index.html. See question 14 regarding cross-reactivity studies related to the SARS-CoV-2 Antibody (IgG), Nucleocapsid, Qualitative immunoassay [test code 39749].

Quest only offers SARS-CoV-2 IgM (spike) in a panel that includes a separate IgG (nucleocapsid) determination.4,5 The results from this qualitative test for SARS-CoV-2 IgM can be positive (reactive) or negative (non-reactive).5,17

A positive (reactive) result indicates that antibodies to SARS-CoV-2 were detected and the individual has potentially been infected with SARS-CoV-2.1,3,5,17 SARS-CoV-2 IgM is generally detectable in blood several days after initial infection. Detection of IgM may indicate a recent infection.1,5,12,13,17 A spike SARS-CoV-2 IgM test may detect an immune response in individuals who have received a COVID-19 vaccination (and who have not been infected naturally), but the clinical significance is unknown.  The clinical significance is unknown, in part, because the performance characteristics of this test in these individuals has not been established.

It is unknown how long SARS-CoV-2 IgM remains detectable, post-infection.1,3,5,17

Whether SARS-CoV-2 antibodies confer immunity to infection also remains unknown.1,3,5,17 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.1

A negative test result means that SARS-CoV-2 IgM was not present in the specimen at levels above the limit of detection.1,5,17 However, patients tested early after infection may not have detectable antibodies despite active infection.1,5,17 A negative result should not be used to rule out infection. Testing of patient specimens for the presence of virus material (eg, RNA or antigen) should be performed if acute infection is suspected.1,3,5,11

A recent publication supports the use of simultaneous IgG/IgM measurements (regardless of method) to significantly improve assay sensitivity.9,10 Regardless of the test result, individuals should continue to follow CDC guidelines to reduce the risk of infection, including social distancing and wearing masks.1

A SARS-CoV-2 semi-quantitative IgG test result is interpretated as a positive value at an index6,7 of ≥1.00. The assay’s positive semi-quantitative reportable range is 1.00 to 20.00. Values reported as ˂1.00 are interpretated as negative. This positive result means that an individual has developed an immune response to recent/prior SARS-CoV-2 infection within the limit of detection of the assay.6,7,15,16 Positive results may also occur after a COVID-19 vaccination, but the clinical significance is not yet known.1,11-13 There are currently no FDA authorized tests for individuals who have received a COVID-19 vaccination, and the performance characteristics of the authorized antibody tests have not been established for these individuals. Since much is still not known about the body’s immune response to this virus, this test result cannot be used to indicate a level of immunity or rule out the chance for reinfection.1,5,14 This test should not be used to diagnose current SARS-CoV-2 infection.1,5,14 If a current infection is suspected, direct molecular or antigen testing for SARS-CoV-2 is necessary.1,3,5-7,15,16

A negative semi-quantitative antibody result means that the patient serum specimen had no SARS-CoV-2 spike IgG antibodies, or that the relative level of antibodies in the patient specimen was below the index cutoff.6,7

Note: this index value should not be interpreted as a measurement of durable immunity or protection from SARS-CoV-2 infection or reinfection.1,3,6,7 Durability and duration of immunity to SARS-CoV-2 are still not clearly defined and continue to be an area of national and global scientific and public health research.1,3 Results obtained with this assay may not be used interchangeably with values obtained with different manufacturers' test methods.1

The result of the total semi-quantitative SARS-CoV-2 antibody immunoassay test is reported as positive at an index of ≥0.8 (index detection interval8 0.4-250.0 U/mL). This positive result means that an individual has developed an immune response to a recent or past infection by SARS-CoV-2 assay.8,18 Positive results may also occur after a COVID-19 vaccination, but the clinical significance is not yet known.1,11-13 At this time, there are no FDA authorized tests for individuals that have received a COVID-19 vaccination and the performance characteristics of the authorized antibody tests have not been established for those individuals. Since much is still not known about the body’s immune response to this virus, this test result cannot be used to indicate a level of immunity or rule out the chance of reinfection.8,18 This test should not be used to diagnose current SARS-CoV-2 infection. If a current infection is suspected, direct molecular or antigen testing for SARS-CoV-2 is necessary.8,18

Conversely, a negative result is reported at an index8 of ˂0.8 U/mL. A negative semi-quantitative antibody result means that the patient serum specimen had no detectable level of SARS-CoV-2 spike IgG antibodies, or that the relative level of antibodies in the patient specimen was below the index cutoff.8,18

Note: this index value should not be interpreted as a measurement of durable immunity or protection from SARS-CoV-2 infection or reinfection.1,3,8 Durability and duration of immunity to SARS-CoV-2 are still not clearly defined and continue to be areas of national and global scientific and public health research.1,3 Results obtained with this assay may not be used interchangeably with values obtained with different manufacturers' test methods.

The presence of IgG and/or IgM antibody to SARS-CoV-2 indicates that the patient has developed an immune response to the SARS-CoV-2 virus. The presence of IgG and/or IgM antibody to SARS-CoV-2 could also indicate potential immune response to a spike vaccine.1,3-5 The clinical significance of a positive antibody result for individuals that have received a COVID-19 vaccine is unknown. The performance of this test has not been established in the context of COVID-19 vaccines. Although the immune response may protect against reinfection, this has yet to be conclusively established.1,3,14,19,20 It is not known how long antibodies to the virus will protect someone, if at all. Cases of reinfection with COVID-19 have been reported but remain rare.1,3,14,19,20

The tests in this panel are intended for qualitative detection of IgG and IgM antibodies to SARS-CoV-2 in human serum and plasma (lithium heparin). The tests are intended for use as an aid in identifying individuals with an adaptive immune response to SARS-CoV-2, indicating recent or prior infection.4,5 At this time, it is unknown for how long antibodies persist following infection and if the presence of antibodies confers protective immunity.1 The test should not be used to diagnose or exclude acute SARS-CoV-2 infection.1 Results are for the detection of SARS-CoV-2 IgG and IgM antibodies. IgG and IgM antibodies to SARS-CoV-2 are generally detectable in blood several days after initial infection, although the duration that antibodies are present post-infection is not well characterized. Individuals may have detectable virus present for several weeks following seroconversion.

Positive results may also occur after a COVID-19 vaccination, but the clinical significance is not yet known.1,11,12,17 At this time, there are no FDA authorized tests for individuals who have received a COVID-19 vaccination, and the performance characteristics of the authorized antibody tests have not been established for those individuals. Clinical researchers continue to investigate what constitutes SARS-CoV-2 protective immunity and long-term vaccine efficacy.11,13,14 This is an area of active clinical research. The CDC states, “Antibody testing is not currently recommended to assess for immunity to COVID-19 following COVID-19 vaccination or to assess the need for vaccination in an unvaccinated person.”1

Manufacturers’ validation of SARS-CoV-2 IgG assays utilized by Quest reported less than 1% positivity in specimens from pre-COVID times4,6,7 (2010, 2017, and 2019).

In an antibody cross-reactivity study of 143 specimens from 30 different categories of infectious disease and autoimmune disease, only 1 specimen (with rheumatoid factor) had a SARS-CoV-2 IgM-positive result.5 In a medical condition cross-reactivity study using 65 specimens from 13 different categories, only 1 specimen (from a hemodialysis patient) tested positive for SARS-CoV-2 IgM.5

The cross-reactivity of the Quest SARS-CoV-2 IgG and SARS-CoV-2 IgM immunoassays (CMIAs)4,6-7 is <1%.

Neutralization assays (NAbs) are performed to help assess the ability of NAbs in patient serum to neutralize virus infectivity in vitro.19 These tests are potentially useful to assess whether past infection (or potential vaccination) has provided protection against infection. The clinical significance of a positive NAb result for individuals who have received a COVID-19 vaccine is unknown. In these tests, serial dilutions of patient serum are incubated with known infectious virus concentrations and then added to virus susceptible cell lines.19 After incubation, infected cells are quantified to determine the effectiveness of patient antibodies in neutralizing the virus.20,21 The presence of NAbs may suggest in vivo immunity to viral infection. Convalescent sera with NAbs are being studied as a treatment for severe COVID-19.19,20 Testing for NAbs is only available at some specially equipped public health laboratories and research facilities. SARS-CoV-2 NAb testing is not performed at Quest.

No, a positive result does not have implications for vaccination. At this time, the CDC and ACIP guidance states that previous SARS-CoV-2 infection, whether symptomatic or asymptomatic, is not considered a contraindication to vaccination, and that serologic testing for SARS-CoV-2 antibodies is not recommended prior to vaccination.1,22

No, semi-quantitative tests can vary significantly, both in the range of detection and, more importantly, in the unit of measure. Not all assays use the same index cutoff for positive and negative; some assays use U/mL and others provide no unit of measure for the reported semi-quantitative index value. Therefore, SARS-CoV-2 semi-quantitative antibody values may not be used interchangeably with values obtained from different manufacturers' test methods. As evidenced in the FDA expanded authorization of semi-quantitative tests that may be used in the determination of high-titer COVID-19 convalescent plasma (CCP) donors, qualifying results of different test methods are expressed with different units and measurements.23

A semi-quantitative index value should not be interpreted as a measurement of durable immunity or protection from SARS-CoV-2 infection or reinfection.1,3,8

References

  1. Interim guidelines for COVID-19 antibody testing. Centers for Disease Control and Prevention. Updated March 17, 2021. Accessed April 7, 2021. https://www.cdc.gov/coronavirus/2019-ncov/lab/resources/antibody-tests-guidelines.html#anchor_1590280017822
  2. FAQs on testing for SARS CoV-2. US Food and Drug Administration. Updated March 25, 2021.  Accessed April 19, 2021. https://www.fda.gov/medical-devices/coronavirus-covid-19-and-medical-devices/faqs-testing-sars-cov-2
  3. EUA authorized serology test performance. US Food and Drug Administration. Updated January 8, 2021. Accessed April 7, 2021. https://www.fda.gov/medical-devices/emergency-situations-medical-devices/eua-authorized-serology-test-performance
  4. Architect SARS-CoV-2 IgG (nucleocapsid). Package insert. Abbott Laboratories; 2020. Accessed April 7, 2021. https://www.fda.gov/media/137383/download
  5. Architect AdviseDx SARS-CoV-2 IgM (spike). Package insert. Abbott Laboratories; 2020. Accessed April 7, 2021. https://www.fda.gov/media/142940/download
  6. Atellica® IM SARS-CoV-2 IgG (COV2G; spike). Instructions for use. Siemens Healthcare Diagnostics Inc; 2020. Accessed April 7, 2021. https://www.fda.gov/media/140699/download
  7. ADVIA® Centaur® SARS-CoV-2 IgG (COV2G; spike). Instructions for use. Siemens Healthcare Diagnostics Inc; 2020. Accessed April 7, 2021. https://www.fda.gov/media/140704/download
  8. Cobas® Elecsys Anti-SARS-CoV-2 S (spike). Instructions for use. Roche Diagnostics Inc; 2021. Accessed April 7, 2021. https://www.fda.gov/media/144037/download
  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;92(9):1518-1524. doi:10.1002/jmv.25727
  11. Livingston EH, Malani PN, Creech CB. The Johnson & Johnson vaccine for COVID-19. JAMA. 2021. doi:10/1001/jama.2021. 2927
  12. Polack FP, Thomas SJ, Kitchin N, et al. Safety and efficacy of the BNT162b2 mRNA Covid-19 vaccine. N Eng J Med. 2020;383(27):2603-2615. doi:10.1056/NEJMoa2034577
  13. Baden LR, El Sahly HM, Essink B, et al. Efficacy and safety of the mRNA-1273 SARS-CoV-2 vaccine. N Eng J Med. 2021;384(5):403-416. doi:10.1056/NEJMoa2035389
  14. Fact sheet for healthcare providers: SARS-CoV-2 IgG (nucleocapsid) assay - Abbott Laboratories Inc. US Food and Drug Administration. Updated December 1, 2020. Accessed April 7, 2021. https://www.fda.gov/media/137381/download
  15. Fact sheet for healthcare providers: Siemens Healthcare Diagnostics Inc Atellica® IM SARS-CoV-2 IgG (COV2G; spike). US Food and Drug Administration. Updated July 31, 2020. Accessed April 7, 2021.  https://www.fda.gov/media/140698/download
  16. Fact sheet for healthcare providers: Siemens Healthcare Diagnostics Inc ADVIA Centaur® SARS-CoV-2 IgG (COV2G; spike). US Food and Drug Administration. Updated July 31, 2020. Accessed April 7, 2021.  https://www.fda.gov/media/140702/download
  17. Fact sheet for healthcare providers: Abbott Laboratories AdviseDx SARS-CoV-2 IgM (spike). US Food and Drug Administration. Updated October 9, 2020. Accessed April 7, 2021. https://www.fda.gov/media/142938/download
  18. Fact sheet for healthcare providers: Roche Diagnostics Inc Elecsys Anti-SARS-CoV-2 S. US Food and Drug Administration. Updated November 25, 2020. Accessed April 7, 2021. https://www.fda.gov/media/144035/download  
  19. Reinfection with COVID-19. Centers for Disease Control and Prevention. Updated October 27, 2020. Accessed April 7, 2021. https://www.cdc.gov/coronavirus/2019-ncov/your-health/reinfection.html
  20. 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
  21. 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
  22. Interim considerations for COVID-19 vaccination of healthcare personnel and long-term care facility residents. Centers for Disease Control and Prevention. Reviewed December 3, 2020. Accessed April 7, 2021. https://www.cdc.gov/vaccines/hcp/acip-recs/vacc-specific/covid-19/clinical-considerations.html
  23. Convalescent plasma EUA letter of authorization. US Food and Drug Administration. Updated March 9, 2021. Accessed April 7, 2021. https://www.fda.gov/media/141477/download

 

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.

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