Hepatitis B – Serology

Consistent with O. Reg. 671/92 of the French Language Services Act, laboratory testing information on this page is only available in English because it is scientific or technical in nature and is for use only by qualified health care providers and not by members of the public.

This page provides hepatitis B virus (HBV) serology testing information at Public Health Ontario (PHO). Information is specific to HBV serology. For information regarding other testing options for HBV, refer to the following PHO webpages:


  • Updated test indications to include hepatitis B virus reactivation (HBVr) and pre-chemotherapy screening
  • Updated minimum sample volume and collection kit
  • Provided a table with HBV serology ordering instructions

Testing Indications

Testing for HBV serology may be indicated in the following:

  • suspected acute or chronic hepatitis
  • pre-natal screening
  • determining the immune status (either following recovery from natural infection or as a result of immunization)
  • prior to starting chemotherapy
  • occupational and other exposures
  • investigation of known or suspected infection control breaches
  • outbreak situations
  • hepatitis B virus reactivation (HBVr) in those who are chronic carriers
  • immune status of at-risk patients to HBVr

If ordering as part of prenatal screening, see Prenatal – Serology.

Acceptance/Rejection Criteria

Donor testing is not available through PHO’s laboratory. Specimens from patients being screened as potential donors (e.g. organ, tissue, cells, fertility, etc.) should be referred to a laboratory that performs donor screening assays. Specimens received for donor screening at PHO’s laboratory will be rejected.

Specimen Collection and Handling

Specimen Requirements

Test Requested Required Requisition(s) Specimen Type Minimum Volume Collection Kit

Hepatitis B Serology


1.5 mL

Vacutainer tubes – Serum Separator Tube (SST); Gold top

Submission and Collection Notes


Complete all fields of the requisition form, including:

  • Test(s) requests (see Special Instructions) and reason for test
  • Patient setting and specimen type and site
  • Relevant clinical information

Label the specimen tube 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. For additional information see: Criteria for Acceptance of Patient Specimens. Failure to provide this information may result in rejection or testing delay.


Refer to instructions for ordering hepatitis B testing for how to complete the “Hepatitis Serology section of the general test requisition.


One FULL 5 mL SST is required for testing a combination of hepatitis virus, HIV, HTLV, syphilis and rubella.


Do NOT submit glass tubes.


Heat inactivated, haemolysed, icteric, lipemic or microbially contaminated serum is not recommended for testing.

Storage and Transport

Centrifuge the specimen tube as per recommended laboratory protocol.

Place specimen tube in biohazard bag and seal. Place completed General Test Requisition in the pouch at the front of the biohazard bag.

Specimen tubes should be stored at 2-8°C following collection and shipped to PHO’s laboratory on ice packs within 3 days of collection.

Specimen tubes received more than 6 days post-collection will not be tested. If delayed shipping is anticipated, remove serum from clot and store frozen at -20°C or colder and ship on dry ice.

All clinical specimens must be shipped in accordance to the Transportation of Dangerous Good Act.  

Special Instructions

HBV serology ordering instructions

Test(s) Requested on Requisition Test(s) Performed Notes
Hepatitis Serology: Chronic Infection HBsAg,
HBcAb Total (IgG+IgM)
Hepatitis Serology:
Acute Infection
HBcAb Total (IgG+IgM)
*HBcIgM will be performed only if HBcAb Total (IgG+IgM) is reactive
Hepatitis Serology: Immune Status Anti-HBs  
Test(s) Requested:
Pre-chemotherapy screening
HBcAb Total (IgG+IgM),
For cancer patients prior to commencing systemic therapy (chemotherapy, targeted therapy, immunotherapy)
Hepatitis Serology: Chronic and Immunity HBsAg,
HBcAb Total (IgG+IgM),
Reason for test must be indicated e.g. Dialysis
Hepatitis Serology:
Acute and Immunity
HBcAb Total (IgG+IgM),
Reason for test must be indicated e.g. Dialysis
*HBcIgM will be performed only if HBcAb Total (IgG+IgM) is reactive
Hepatitis Serology: Chronic Infection and
Test(s) Requested: HBeAg, HBeAb
HBcAb Total (IgG+IgM)
Testing for HBeAg and HBeAb will only be performed if HBsAg is reactive
Hepatitis Serology:
Acute Infection and
Test(s) Requested: HBeAg, HBeAb
HBcAb Total (IgG+IgM)
Testing for HBeAg and HBeAb will only be performed if HBsAg is reactive
Test(s) Requested: HBsAg,
Do not complete the Hepatitis Serology section.
Under ‘Reason for Test’, select ‘Other’ and specify Hepatitis B Exposure
Test(s) Requested:
HBeAg and HBeAb
HBeAg and HBeAb
Do not complete the Hepatitis Serology section.
Enter the specific test(s) under Test(s) Requested on the General Test Requisition
Needlestick/Occupational exposure (Source Patient) HBsAg Refer to STAT and Critical Specimens Testing below
Needlestick/Occupational exposure (Exposed Patient) HBsAg

Requisitions and Kit Ordering

Test Frequency and Turnaround Time (TAT)

HBV serology testing is performed daily Monday to Friday.

Turnaround time may be up to 3 business days from receipt of the specimen by PHO’s laboratory for “non-reactive” specimens and up to 6 business days for “reactive” specimens.

STAT and Critical Samples Testing

If warranted, testing of the ‘source patient’ for needle stick/occupational exposures and high risk patients presenting in labour who have not received prenatal care or for whom results of previous testing are not readily available are expedited.

Prior to sending, please notify PHO’s laboratory Customer Service Centre at 416-235-6556/1-877-604-4567 immediately. Contact the After-Hours Emergency Duty Officer at 416-605-3113 if outside of PHO’s laboratory operating hours.

STAT specimens must be shipped separately from routine specimens, the outer package is clearly marked indicating ‘STAT’. It must be 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.

Test Methods

Specimens submitted for HBV serology are tested using a chemiluminescent microparticle immunoassay (CMIA) for the qualitative detection of HBV markers (either antibodies or antigens).


If HBsAg is reactive, additional testing for HBcAb Total (IgG + IgM) will be performed automatically.

If HBsAg is reactive, but HBcAb Total is non-reactive then the specimen will be tested using an HBsAg confirmatory test (‘blocking assay’). Other markers are tested based on the test request as described under Special Instructions.


Please refer to CDC Resources for interpretation of HBV serology test results.


Results are reported to the ordering physician, authorized health care provider (General O. Reg 45/22, s.18), or submitter as indicated on the requisition.

Specimens that are positive for HBV surface antigen (HBsAg) are reported to the Medical Officer of Health as per the Ontario Health Protection and Promotion Act.

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Updated 13 April 2023