Ultrasenstive Immunoassay For SIV P27
AIDS RESEARCH AND HUMAN RETROVIRUSES | JUNE 05, 2018
Swanstrom AE, Gorelick RJ, Wu G, Howell B, Vijayagopalan A, Shoemaker R, Oswald K, Datta SA, Keele B, Del Prete G, Chertova E, Bess J, Jr. and Lifson J
AIDS Research and Human Retroviruses
Although effective at suppressing viral replication, combination antiretroviral treatment (cART) does not represent definitive therapy of HIV infection due to persistence of replication-competent viral reservoirs. The advent of effective cART regimens for simian immunodeficiency virus (SIV) infected nonhuman primates (NHP) has enabled the development of relevant models for studying viral reservoirs and intervention strategies targeting them. Viral reservoir measurements are crucial for such studies, but are problematic. Quantitative PCR (qPCR) overestimates the size of the replication competent viral reservoir, as not all detected viral genomes are intact. Quantitative viral outgrowth assays (QVOA) measure replication competence, but suffer from limited precision and dynamic range, and large numbers of cells. Ex vivo virus induction assays to detect cells harboring inducible virus represent an experimental middle ground, but detection of inducible viral RNA in such assays does not necessarily indicate production of virus, while detection of more immunologically relevant viral proteins, including p27CA, by conventional enzyme-linked immunoabsorbent assays (ELISA) lacks sensitivity. We have developed an ultrasensitive digital SIV Gag p27 assay, which is 100-fold more sensitive than a conventional ELISA. We show that in ex vivo virus induction assays the quantification of SIV Gag p27 produced by stimulated CD4+ T cells from rhesus macaques receiving cART enabled earlier and more sensitive detection than conventional ELISA-based approaches and was highly correlated with SIV RNA, as measured by quantitative reverse transcription PCR (qRT-PCR). This ultrasensitive p27 assay provides a new tool to assess ongoing replication and reactivation of infectious virus from reservoirs in SIV-infected NHP.
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