New 5-Minute Test Accurately Distinguishes HIV Infection from Vaccine-Induced False Positives

Researchers have developed a new rapid diagnostic test that delivers results in 5 minutes, accurately distinguishing an active HIV-1 infection from a vaccine-induced false positive. The device overcomes vaccine-induced seropositivity (VISP), a major roadblock in HIV vaccine development where standard tests cannot differentiate between antibodies from a vaccine and those from a true infection. By simultaneously detecting both protein and nucleic acid markers in a single plasma sample, the new 3D-printed device provides a definitive result.

In a clinical study using 104 clinical plasma samples, the test demonstrated 95% sensitivity in identifying active HIV-1 infection and 98% specificity in identifying cases with vaccine-induced molecules. The findings were published on December 3, 2025, in Science Advances.

The Problem: The Clinical Challenge of Vaccine-Induced Seropositivity (VISP)

Vaccine-induced seropositivity presents a significant obstacle in HIV vaccine research. The phenomenon occurs because HIV vaccines are designed to elicit the production of antibodies that target key viral proteins—the same proteins that standard serologic tests detect to confirm an infection. Consequently, vaccinated individuals who are not infected can produce a false-positive test result. The scope of this problem is substantial, with VISP occurring in anywhere from 0.4% to over 95% of tests in vaccine trials, depending on the vaccine design, diagnostic tests used, and patient demographics.

“A direct consequence of this immunogenic overlap is that the vaccine can make a person test positive for HIV-1, even when they do not have the infection,” corresponding author Dipanjan Pan, PhD, Dorothy Foehr Huck & J. Lloyd Huck Chair Professor in Nanomedicine, Department of Materials Science and Engineering, Pennsylvania State University, said in a press release. “This poses serious social, professional and personal consequences for those individuals. It can also make trial results difficult to interpret, slowing the progress toward widely available HIV vaccines.”²

Individuals with VISP can be disqualified from donating blood, bone marrow, or other organs and may face complications with insurance, military service, employment, travel, immigration, or pregnancy.

“VISP and seroreactivity are not just technical nuisances; they can lead to ongoing misdiagnosis with broad effects on individuals and populations, such as psychological distress and societal challenges, including misunderstanding within families and communities,” Pan noted.² This complication hinders trial recruitment and forces current diagnostic procedures to rely on costly and complex nucleic acid amplification tests (NAATs), which require specialized equipment and personnel, with results often taking hours or days to return. The new test’s five-minute turnaround starkly contrasts with these time-consuming methods.

The Solution: A Multiplexed Electrochemical Assay

To solve the VISP challenge, the research team developed a 3D-printed device that uses a multiplexed electrochemical assay to analyze a plasma sample. The platform simultaneously tests for three distinct biomarkers, providing a comprehensive diagnostic picture that can separate a vaccine-induced immune response from an active viral infection.

The three key biomarkers it detects are:

1. p24 antigen (a core viral protein)
2. anti-p24 antibody (the antibody produced by the immune system in response to the p24 protein)
3. HIV-1 viral RNA (the virus's genetic material)

The primary innovation lies in its ability to detect the virus's genetic material alongside protein markers.

"By incorporating HIV-1 RNA detection, the testing platform provides a definitive indicator of active viral replication, which is absent in VISP cases. As such, this test can accurately discriminate between vaccine-induced responses and true infection," Pan stated in the press release. By shifting the diagnostic paradigm from a single-target antibody search to a multi-faceted hunt for both proteins and active viral RNA, the device eliminates the ambiguity that has plagued HIV diagnostics in vaccine trials for decades.²

Clinical Validation and Performance Metrics

The new test was validated using 104 clinical plasma samples obtained from the HIV Vaccine Trials Network (HVTN). The samples represented four distinct groups: HIV-negative placebo, HIV-negative vaccinated, HIV-positive placebo, and HIV-positive vaccinated. The platform demonstrated high accuracy, achieving 95% sensitivity in identifying active HIV-1 infection and 98% specificity in identifying cases where only vaccine-induced molecules were present.

The test's analytical limits of detection (LOD) were quantified for each biomarker:

• p24 antigen: 5.88 picograms per milliliter
• anti-p24 antibody: 10.96 picograms per milliliter
• HIV-1 RNA: 1259 copies per milliliter

Diagnostic accuracy was further confirmed through Receiver Operating Characteristic (ROC) analysis, which measures a test's ability to distinguish between groups. The Area Under the Curve (AUC) values for each marker underscored the test's strong discriminatory power:

• HIV-1 RNA: 0.9888
• anti-p24 antibody: 0.9705
• p24 antigen: 0.9356 These values, all approaching the ideal score of 1.0, indicate exceptional diagnostic accuracy and a high degree of reliability in distinguishing between different patient groups.

Future Implications for Vaccine Trials and Patient Care

The development of this rapid, accurate, and low-cost diagnostic tool has profound implications for the future of HIV research and care. By reliably differentiating VISP from true infection, the test has the potential to accelerate vaccine development, reduce misdiagnosis and the associated psychological distress for trial participants, and improve recruitment for crucial clinical trials. Crucially, the platform adheres to the World Health Organization’s ASSURED criteria for point-of-care diagnostics: Affordable, Sensitive, Specific, User-friendly, Rapid and Robust, Equipment-free, and Deliverable to those who need it. This makes it a viable solution for both well-resourced and resource-limited settings.

Summarizing the platform's value, Pan stated in the press release “Our proposed all-in-one testing platform represents a substantial advancement in HIV diagnostics, enabling accurate detection of active HIV infection while minimizing false positives due to VISP.” The research team's next steps include refining the prototype to make it more durable, extending its screening capabilities to other pathogens, and exploring its potential as an at-home viral load test for patients undergoing antiretroviral therapy.^1,2

References:

1. Dighe K, Colak O, Moitra P, et al. Distinguishing active HIV-1 infection from vaccine-induced seropositivity in HIV vaccine trial participants. Sci Adv. Published online December 3, 2025. doi:10.1126/sciadv.adz5639
2. New test distinguishes vaccine-induced false positives from active HIV infection. News release. Ashley Wenners Herron. December 3, 2025. Accessed December 9, 2025. https://www.psu.edu/news/research/story/new-test-distinguishes-vaccine-induced-false-positives-active-hiv-infection