Hepatitis B is a viral infection that targets the liver, and early identification is important for patient care and public health. Screening tests are routinely used to detect the virus, but a positive result can cause significant worry, especially when the result is inaccurate. Understanding the factors that lead to a false-positive test result is necessary for patients and providers to navigate the diagnostic process. False positives occur when the screening process mistakenly identifies the presence of the virus, requiring careful follow-up to determine the true infection status.
How Hepatitis B Screening Tests Work
Initial screening for Hepatitis B infection typically uses a blood test designed to detect the Hepatitis B Surface Antigen (HBsAg). This antigen is a protein on the outer surface of the virus, and its presence indicates an active infection. The laboratory technique employed is usually a highly sensitive immunoassay, such as a chemiluminescent microparticle immunoassay.
These assays use manufactured antibodies specifically designed to bind to the HBsAg protein in a patient’s blood sample. If the viral protein is present, it forms a complex with the test antibodies, triggering a measurable signal, often light emission. A signal exceeding a predetermined cutoff value is reported as “reactive” or “positive,” indicating a marker was detected, but not necessarily establishing a true, active Hepatitis B infection.
Immunological Cross-Reactivity
Immunological cross-reactivity is the most common cause of false-positive results. This occurs when the test antibodies designed to capture HBsAg mistakenly bind to a different, non-HBV substance circulating in the bloodstream. The test kit interprets this binding as a true positive because the binding mechanism mirrors that of the target antigen.
Interference from Infections and Autoimmunity
A recent, non-HBV viral infection can stimulate the immune system to produce antibodies structurally similar to the viral antigen. Infections such as Epstein-Barr virus, Parvovirus B19, or acute hepatitis A have been associated with this broad immune system activation. These non-specific circulating antibodies bind to the test reagents, generating a false signal.
Autoimmune conditions can also introduce interfering substances. For example, individuals with Rheumatoid Arthritis may have high levels of Rheumatoid Factor. This autoantibody can bind non-specifically to the mouse monoclonal antibodies used in many commercial test kits, mimicking the HBsAg-antibody complex and leading to an inaccurate reading.
Vaccine and Therapeutic Interference
Recent vaccinations, even those not against Hepatitis B, can trigger a temporary immune response that causes cross-reactivity. The stimulation of the immune system by a vaccine, such as the influenza or rabies shot, can lead to the transient production of antibodies that interfere with the HBsAg assay. Furthermore, the Hepatitis B vaccine itself contains the recombinant HBsAg protein. Testing performed shortly after administration can detect this vaccine component, causing a temporary false-positive result.
Another biological interference comes from heterophile antibodies, which are produced in response to general immune system challenges. These antibodies can bridge the gap between the test’s capture and detector antibodies, creating a false-positive signal without HBsAg being present. Certain therapeutic agents, such as granulocyte-colony stimulating factor, have also been implicated in transient false-positive HBsAg results.
Technical and Procedural Errors
Non-biological factors related to the lab environment and testing process can also produce inaccurate positive results. Screening assays are designed for high sensitivity, meaning they detect minute amounts of HBsAg. However, this high sensitivity can sometimes reduce specificity, increasing the likelihood of background noise being read as positive. While the assay’s cutoff value attempts to balance this trade-off, marginal results remain challenging.
Issues within the laboratory can compromise the integrity of the sample or the reagents. Potential sources of error include:
- Improper handling of the blood sample, such as incorrect storage temperatures or contamination during processing.
- Reagent deterioration due to improper storage, expiry, or variability between manufacturing lots.
- Operator error, including mislabeling samples, incorrect data entry, or errors in interpreting results close to the cutoff point.
- Reagent carryover in automated testing, where trace amounts from a highly positive sample contaminate the equipment used for a subsequent sample.
Confirming or Dismissing a Positive Result
An initial positive HBsAg screening result requires follow-up with more specific, high-reliability tests. A reactive result is not a final diagnosis but a trigger for investigation to distinguish a true infection from a false positive.
The first step in confirmation is typically the HBsAg neutralization assay, which proves the detected substance is the Hepatitis B surface antigen. In this test, the patient’s sample is mixed with a known anti-HBs antibody. If HBsAg is truly present, the antibody neutralizes it, causing the immunoassay signal to drop significantly (often by more than 50 percent).
If the signal fails to drop after neutralization, it confirms the initial positive result was due to a non-specific cross-reacting substance, dismissing the Hepatitis B diagnosis. For samples confirmed positive by neutralization, the next step is often molecular testing, such as Polymerase Chain Reaction (PCR). PCR directly detects and quantifies the Hepatitis B viral DNA, providing definitive evidence of an active infection.