MRC-University of Glasgow Centre for Virus Research

New research from the MRC-University of Glasgow Centre for Virus Research has uncovered a flaw in widely used virus immunity tests. It turns out that HIV drugs can interfere with lab results, leading to false positives that throw off our understanding of immune responses. This is especially problematic in global health studies where HIV prevalence is high.

The discovery began when researchers noticed something odd across two separate studies. One study looked at immune responses to SARS-CoV-2 in a population-based Malawian cohort, and the other studying hepatitis C virus (HCV) in acute HCV infected participants from the UK. Despite their different settings, both populations had a high prevalence of HIV, and both studies used a well-known serology method known as a pseudotyped virus neutralisation assay (PVNA).

These assays are a standard method in vaccine efficacy studies and virus surveillance. Scientists create modified viruses that can’t replicate, then dress them up with the outer protein of whatever virus they’re studying – like putting a SARS-CoV-2 costume on a harmless virus. The SARS-CoV-2 assay used a HIV-based pseudovirus system (Malawi cohort), while the HCV assay used a murine leukaemia virus (MLV)-based pseudovirus system (UK cohort).

However, something was clearly wrong with the result.  

People living with HIV typically show weaker immune responses due to their compromised immune system. Instead, these participants showed unusually high levels of neutralising activity. This was the complete opposite of what the researchers expected to see and therefore concerns were raised about the accuracy of the tests for these people living with HIV. The researchers then decided to dig deeper.

Researchers turned to a different assay system based on vesicular stomatitis virus (VSV), which isn’t related to HIV or MLV. The results from the VSV-based assays finally made sense and people living with HIV were showing lower neutralising activity. Live virus tests supported this finding, confirming that something was artificially inflating the results in the original retroviral based tests.

To test this, the researchers ran a control assay using a retroviral system with a different target protein, VSV-glycoprotein. Once again, the neutralising activity in people living with HIV was incredibly high, even though the participants had no known exposure to VSV. The culprit had to be the retroviral backbone itself.

The breakthrough came when researchers looked at medications being taken by study participants—specifically antiretroviral therapy (ART). In the UK study, where detailed medication data was available, a clear pattern emerged: people taking integrase inhibitors (a widely used class of HIV drugs) were more likely to show interference in the MLV-based HCV assay.

Though the Malawi study lacked detailed ART data, previous research hinted to integrase inhibitors potentially interfering with HIV-based SARS-CoV-2 pseudovirus systems (De La Torre-Tarazona et al. 2023)—like the one used with the Malawi cohort. To prove this, the team tested three common antiretroviral drugs directly against the pseudovirus systems in the lab. Dolutegravir, an integrase inhibitor, disrupted both the HIV- and MLV-based assays.

The final piece of evidence came when researchers purified IgG antibodies from participants’ blood samples and tested them separately. The false signals disappeared, confirming that residual drug in the samples, not the antibodies, was the source of the interference.

This discovery has serious implications for viral immunity research worldwide. Retrovirus-based neutralisation assays are commonly used in immunity studies and vaccine trials, including in regions of high HIV prevalence. If these tests are vulnerable to interference from integrase inhibitors, then past results may have overestimated levels of virus-neutralising antibodies.

The path forward requires a fundamental shift in how we measure virus immunity in studies involving people living with HIV and antiretroviral therapy. Researchers are urged to switch to non-retroviral systems like VSV-based assays when working with blood samples from HIV prevalent communities. These systems appear robust against drug interference and are more consistent with live virus testing results. It’s also vital for researchers to collect detailed information about participants’ medication, and to interpret antibody results with caution in regions where HIV is common or where ART use may be underreported.

This research reveals a concerning truth: the very drugs that help people with HIV live healthier lives can inadvertently skew the tests we use to understand their immune responses. Now that we know integrase inhibitors can throw off retrovirus-based assays, the scientific and clinical communities can take concrete steps to make future studies more accurate and reliable.

The implications stretch beyond the laboratory—this could reshape how we design virus surveillance studies, evaluate vaccines, and ultimately understand immunity in one of the world's most vulnerable populations.

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First published: 5 August 2025