NEW YORK – Researchers at Florida Atlantic University are progressing on a project to create a highly sensitive molecular diagnostic system for HIV self-testing. With their recent Phase II funding only slightly delayed by new National Institutes of Health funding policies, the team expects to soon prototype its disposable microfluidic device on clinical samples as part of its path toward commercialization.
Waseem Asghar, a biomedical engineer at FAU and principal investigator on the project, said in an interview that, to his knowledge, there is no molecular HIV self-test currently commercially available in the US or elsewhere. His group hopes to change that with an RT-LAMP testing platform.
The team was granted a two-year Phase II award of $1.3 million from the NIH's National Institute of Allergy and Infectious Diseases division to assess the accuracy of a prototype device with clinical HIV samples collected by collaborators at Emory University and the University of Virginia.
Viral load can be astronomical in the first few weeks following an HIV infection, until the body mounts an immune response, and can rise again in cases where patients are on suppressive medication and acquire treatment resistance.
Rapid lateral flow assays detecting HIV antibodies are most effective about three weeks after infection, while viral antigen tests can be used slightly earlier. Tests that detect viral RNA, on the other hand, can be extremely sensitive, in part because they amplify these relatively rare targets.
Nitika Pant Pai, an HIV researcher at McGill University in Montreal who was not involved with the FAU research, said that antigen-based p24 tests are relatively inexpensive and can detect HIV infection within a window between about 18 days and 45 days after exposure, while antibody-based tests can be used between about 23 days to 90 days after exposure.
However, molecular detection with nucleic acid amplification RNA tests "is the key to identifying infection early," she said, and is effective within a window of between 10 days and 33 days.
Early detection can prevent the spread of HIV during a period when viral load is high and can also connect patients to care sooner, which has potential benefits to their long-term health.
Unfortunately, unlike antibodies — which can be found in saliva and other samples — HIV viral RNA is almost exclusively found in blood. And therein lies a major problem for self-test developers.
Asghar said that with more than 1 billion red blood cells in a milliliter of human blood, the virus is miniscule and rare by comparison. Centrifugation is required to isolate the virus for testing in the lab, he said, but very few people have lab-grade centrifuges in their homes.
The FAU team's solution involves magnetic actuation within the microfluidic chip, such that the test can be sample-in, answer-out, requiring very minimal user manipulation.
The issue of centrifugation has also been noted in an HIV Medicine study published earlier this year. An assay from Cepheid called Xpert HIV VL showed great promise, reviewers said, but its use at the point of care in rural areas was hampered by the need for centrifugation. They also noted that centralized lab tests are expensive and results turnaround can take up to two months in rural areas of some countries.
Portable platforms are also expensive, McGill's Pai said, adding that to her knowledge there are no point-of-care single-use devices available for molecular HIV testing.
"It is the next frontier in testing for sure," she said. However, from her perspective, it is critical to determine assay performance in the context of anti-retroviral drug treatment and to make sure the limit of detection is low.
"The proof of this test will be realized when it is evaluated in a field setting under various temperature conditions," Pai said.
The FAU assay is expected to cost approximately $5 per test. Asghar and his colleagues validated their test system development last year in a study published in Biosensors, describing an automated RT-LAMP-based microfluidic chip that combines the RNA isolation, purification, and amplification steps with a visual readout.
And, in a study published last year in Viruses, the team showed it could detect SARS-CoV-2 within 40 minutes from saliva and nasopharyngeal samples. The team has also previously shown its approach could be used to detect Zika and hepatitis C viruses.
Self-testing is a critical piece of HIV prevention, control, and patient management. The only self-test approved by the US Food and Drug Administration currently is an immunoassay approved in 2012 from OraSure.
Pai and her colleagues recently described an app called HIVSmart! that could enhance the accuracy of HIV self-tests like OraSure's. However, she said her team isn't currently in discussions with industry to integrate the app, although it is keen for it to be taken up by governmental agencies scaling HIV testing globally.
Other HIV molecular self-tests are also potentially on the horizon. For example, a research team at Pennsylvania State University has developed a point-of-care testing system using RT-LAMP that is also being developed for HIV viral load monitoring from whole blood. This team also developed a novel de-clogging mechanism that enables them to adapt a glass nanopore device to the native buffers required for point-of-care RT-LAMP testing.
Still, although a review published in February was a recent example of the consensus that accurate POC HIV RNA tests "could leapfrog fourth-generation POC assays," these tests remain unavailable for routine use.
Meanwhile, there are a handful of point-of-care or near-patient molecular diagnostic test systems for HIV viral load monitoring either commercially available or in development, according to a 2019 evaluation in Clinical Microbiology Reviews. These included Abbott's m-PIMA, formerly known as the Alere q HIV-1/2 Detect, Danaher's Cepheid Xpert HIV-1 Viral Load, the SAMBA I PSQ and SAMBA II WBSQ from Diagnostics for the Real World, and the TrueNat HIV Viral Load from MolBio Diagnostics.
Global health HIV strategies through 2030 developed by the World Health Organization include encouraging genomic approaches — especially those that can be applied to other pathogens — as well as self-testing options that can take HIV testing and patient monitoring to communities and remote areas.
HIV testing cost is a critical barrier in low- and middle-income countries, however. According to a survey by researchers at Johns Hopkins University published in 2023, this is also increasingly true for high-income countries. The survey also revealed that test sensitivity was more important than cost for practitioners in LMICs as well as HICs, although both groups said they would compromise on sensitivity if it would reduce cost. "Low cost should remain a strong consideration during assay development and may be critical to adoption and access in all settings," the authors concluded.
In addition to qualitative HIV testing, the FAU team aims to develop a quantitative test in the future and is also adapting its system for other infectious diseases, Asghar said. The team currently has a preliminary patent on the system and hopes to commercialize it through collaboration with industry.