Coronavirus Disease 2019 (COVID-19) – Serology

This test provides qualitative detection of serum IgG antibodies to SARS-CoV-2, the causative agent of novel coronavirus disease (COVID-19).

Due to the potential for false negative and false positive results, COVID-19 serology should not be used as a diagnostic test except in very rare circumstances. SARS-CoV-2 antibodies do not correlate with recovery or infectivity. Serology also cannot be used to assess whether a person is immune to COVID-19 or to determine their COVID-19 vaccination status.

* Copie d'impression non contrôlée. Valide uniquement le jour de l'impression : 01 juin 2023.

Testing Indications

This test is intended for use as an aid in determining if a patient with an adaptive immune response has been previously exposed to SARS-CoV-2. Currently, it is not known whether the presence of SARS-CoV-2 IgG antibodies correlates with immunity. It can take at least 7-14 days from symptom onset to develop a measurable SARS-CoV-2 IgG response with some individuals never seroconverting. In evaluations at Public Health Ontario (PHO), the highest sensitivity of the assay was seen at >14-21 days from symptom onset. Further, the duration of the IgG response is variable, with a reduction in IgG levels and seronegativity in as little as 2-3 months in some patients. Additional information about COVID-19 serology testing can be found here: What We Know So Far – COVID-19 and Serology Testing.

In patients with a low pretest probability (e.g. no high-risk exposure or symptoms compatible with COVID-19), there is a risk of a false positive COVID-19 serology result, even with high test specificity. Conversely, there is an increased risk of a falsely negative interpretation if serum is taken when antibody levels are too low to be detected, such as when it is too early (i.e., <2-3 weeks after symptom onset) or too late (i.e. antibody waning), from a patient with a mild infection, or if the patient is immunocompromised. Given these caveats and significant gaps in our understanding of the immune response in COVID-19, serology testing has limited clinical value for individual patients. Detection of viral RNA by molecular testing, such as PCR, is the gold standard for diagnosing COVID-19 in suspected patients. If the patient is symptomatic and is eligible for diagnostic laboratory testing according to the current testing guidelines, a respiratory specimen should be tested by COVID-19 PCR Testing.

Currently, the limited clinical value for individual patient testing precludes the widespread use of COVID-19 serology as a clinical diagnostic tool. It may be considered for clinical use as an adjunct to COVID-19 PCR testing in:

Patients suspected to have multisystem inflammatory syndrome in children (MIS-C) or adults (MIS-A) with a negative, indeterminate, or inconclusive PCR test result or who were not tested
Treatment decisions for patients who are eligible for monoclonal antibody therapy as outlined in the Science Table Brief on Evidence-Based Recommendations for Use of Casirivimab + Imdevimab, and Sotrovimab for Adults in Ontario. Specifically, serology testing is recommended for moderately/critically ill patients who are within 9 days of symptom onset:

and are clinically stable with no history of COVID-19 or full vaccination; OR
have a history of COVID-19 or are fully vaccinated

**Specimens submitted for monoclonal antibody treatment decision making MUST be clearly marked as “Priority – for monoclonal antibody treatment” and sent following the instructions described under Special Instructions. Failure to do so may result in testing delays.

Other clinical scenarios of severe illness with negative PCR tests, where serology results may be helpful for clinical management and/or public health action, will be considered following consultation and approval by a PHO Microbiologist before specimen collection. Specimens submitted for testing for indications other than MIS-C/MIS-A, to guide treatment decisions for monoclonal antibody therapy, or without prior approval will be rejected.

COVID-19 serology testing should not be used in an individual who has received a COVID-19 vaccination, outside of the specific indication above, as the test cannot always differentiate between antibodies produced in response to previous infection and vaccination.

Serology should NOT be used for:

the diagnosis of acute infection, reinfection, or determining the infectivity of the patient
determining immune status of the patient (i.e. protection against future infection)
determining COVID-19 vaccination status of the patient or serological response to vaccination

A PHO Microbiologist can be contacted through the PHO's Customer Service Centre at 416-235-6556 / 1-877-604-4567 or customerservicecentre@oahpp.ca.

Specimen Collection and Handling

Specimen Requirements

Test Requested	Required Requisition(s)	Specimen Type	Minimum Volume	Collection Kit
COVID-19 IgG	Coronavirus Disease 2019 (COVID-19) Serology Test Requisition	Serum	1.0 ml	Serum Separator Tubes (SST)

Submission and Collection Notes

Specimens submitted for testing for indications other than the indications as described above, or without prior approval by a PHO Microbiologist will be rejected.

OHIP or other non-PHO requisitions will not be accepted for specimen collection or submission. The dedicated PHO laboratory COVID-19 Serology Test Requisition must be used and submitted.

Complete all fields of the dedicated COVID-19 Serology Test Requisition form (do not use the COVID-19 Virus Test Requisition that is used for PCR testing). Include the patient's full name, date of birth, Health Card Number (must match the specimen label), patient address, physician name and address, specimen type and date of collection, reason for test, patient setting, clinical symptoms (including onset date) and medical history (including COVID-19 vaccination status), COVID-19 PCR result(s) and date(s) of testing, travel, and other exposure history.

PHO prefers centrifuged serum separator tubes for serological assays Place specimen in biohazard bag and seal. Specimens should be stored at 2-8°C following collection and shipped to PHO’s laboratory on ice packs as soon as possible and within 7 days from collection.

Specimens may be stored on or off the clot or red blood cells for up to 1 day at 15-30°C or 7 days if refrigerated at 2-8°C. Stability past 7 days from the collection has not been evaluated at PHO’s laboratory. If shipping/testing will be delayed beyond the recommended storage time, remove serum from the clot and store at -20°C or colder and ship frozen.

Specimens should not be heat-inactivated or pooled. Grossly haemolysed, icteric, lipemic or microbially contaminated sera and specimens with fungal growth will not be tested.

Do NOT submit glass tubes.

Timing of Specimen Collection

Collect serum at least 14-21 days following symptom onset. If collecting earlier, consider collecting paired sera with the second collection at least 14-21 days following symptom onset and within 4 weeks of the first collection (at least 1 week apart). For patients suspected to have MIS-C/MIS-A or if serology testing is being used to guide monoclonal antibody therapy, sera may be collected at the time of investigation.

Storage and Transport

Label the specimen container with the patient's full name, date of collection and one other unique identifier such as the patient's date of birth or Health Card Number. Failure to provide this information may result in rejection or testing delay.

Special Instructions

“PRIORITY” testing is only available to support treatment decisions for patients who are eligible for monoclonal antibody therapy and where serology testing is recommended (see indications above):

Notify PHO PRIOR to sending a “Priority” specimen by contacting our Customer Service Centre at 416-235-6556 / 1-877-604-4567. Failure to do so will result in testing delays.
“Priority” testing is only available Monday to Saturday (not available on Sundays and holidays). The target turnaround time is 24 hours from receipt at PHO’s laboratory Toronto site. Specimens that are received by 1 p.m. Monday to Friday and by 12 p.m. Saturday will be tested on the same day, otherwise the specimens will be tested on the next business day. Specimens that are received at PHO’s laboratory (Toronto) after 12 p.m. on Saturday will be tested on Monday or the next regular business day.
“Priority” specimens must be shipped separately from routine specimens in a clearly marked package indicating “Priority – for monoclonal antibody treatment”, handled in accordance with the Canadian Biosafety Standards, and shipped in accordance with the Transportation of Dangerous Goods Regulations. Failure to ship separately will delay testing. On the COVID-19 Serology Test Requisition, please choose “Priority – for monoclonal antibody treatment” under Reason for Test.
“Priority” specimens must be sent directly to PHO’s laboratory Toronto site to the shipping and receiving dock at 661 University Ave., Toronto, Ontario. For delivery instructions please see Directions to 661 University Shipping Dock for Clinical Samples.

Requisitions and Kit Ordering

Test Frequency and Turnaround Time (TAT)

Routine SARS-CoV-2 IgG serology is performed two days a week. Turnaround time is up to 5 days from receipt by PHO’s laboratory.

“Priority” testing is only available Monday to Saturday (testing is not available on Sundays and holidays). The target turnaround time is 24 hours from receipt at PHO’s laboratory Toronto site. Specimens that are received by 1 p.m. Monday to Friday and by 12 p.m. Saturday will be tested on the same day, otherwise the specimens will be tested on the next business day. Specimens that are received at PHO’s laboratory (Toronto) after 12 p.m. on Saturday will be tested on Monday or the next regular business day.

Test Methods

PHO’s laboratory uses two SARS-CoV-2 IgG assays: the Abbott Alinity SARS-CoV-2 IgG assay that detects anti-nucleocapsid (N) IgG, and the Abbott ALINITY AdviseDx SARS-CoV-2 IgG II assay that detects anti-spike (S) IgG Both are chemiluminescent microparticle immunoassay (CMIA) assays intended for the qualitative detection of IgG antibodies to SARS-CoV-2 in human serum.

Algorithm

Specimens are tested using both assays and each result is reported individually. A reactive result on either assay is considered an overall “Detected” result for SARS-CoV-2 IgG antibody.

Interpretation

SARS-CoV-2 IgG testing should NOT be used to determine a patient's immune status, vaccination status, or infectivity. These assays cannot definitively differentiate between antibodies generated from previous infection and vaccination. Results should be interpreted in the context of clinical and exposure history.

Anti-N IgG Result	Anti-S IgG Result	Interpretation
Non-reactive	Non-reactive	The absence of IgG does not rule out current or previous SARS-CoV-2 exposure or COVID-19 vaccination. If clinical suspicion is high, consider retesting in 2-4 weeks to document seroconversion Negative results may occur if the specimen is collected too soon or too late following infection, if the patient is immunocompromised, or if the patient is too young to produce an effective adaptive immune response (eg. neonate)
Non-reactive	Reactive	Detection of anti-S IgG alone may indicate previous or current exposure to SARS-CoV-2 and/or COVID-19 vaccination A negative anti-N IgG result does not always rule out previous SARS-CoV-2 infection as anti-N IgG may wane faster than anti-S IgG following infection
Reactive	Non-reactive	Detection of anti-N IgG alone may indicate previous or current exposure to SARS-CoV-2 A negative anti-S IgG result does not rule out antibody development from previous COVID-19 vaccination
Reactive	Reactive	Detection of both anti-N and anti-S IgG may indicate previous or current exposure to SARS-CoV-2 or, previous exposure and COVID-19 vaccination

Further information on a reactive SARS-CoV-2 IgG result:

An individual with evidence of seroconversion within a 4-week interval between collections, regardless of nucleic acid amplification testing (e.g. real-time PCR) and in the absence of COVID-19 vaccination, , is considered a "confirmed" case.¹
An individual with antibody detected in a single serum specimen is considered a "probable" case if they had symptoms compatible with COVID-19 AND had a high-risk exposure or epidemiological link AND antibody was detected within four weeks following symptom onset.1
False-positive results may occur from cross-reaction due to prior infection with other human coronaviruses, including SARS-CoV-1 and certain seasonal coronaviruses (e.g. human coronavirus OC43).
In infants less than 12 months old, detected IgG antibody may represent maternal transfer of antibodies.

An invalid SARS-CoV-2 IgG test result:

Indicates that testing could not be completed for technical reasons. This may be due to inadequate specimen integrity. Submit another specimen if clinically indicated.

If the patient is symptomatic and is eligible for diagnostic laboratory testing according to the current testing guidelines, a respiratory specimen should be collected and tested for SARS-CoV-2 using a molecular assay. Additional information can be found here: COVID-19 PCR Testing.

¹Refer to the Ministry of Health COVID-19 Case Definition: https://www.health.gov.on.ca/en/pro/programs/publichealth/coronavirus/docs/2019_case_definition.pdf

Reporting

Results are reported to the ordering physician or health care provider as indicated on the requisition.

As a disease of public health significance, all reactive results will be reported to the local public health unit.