Development of a sensitive, quantitative assay with broad subtype specificity for detection of total HIV-1 nucleic acids in plasma and PBMC.

ABSTRACT: An LTR-based quantitative PCR (qPCR) assay was modified and optimized for the quantification of total HIV-1 nucleic acids in plasma and PBMC. TaqMan qPCR primers and probes were designed against the NCBI/LANL HIV-1 compendium database by analyzing sequences used in assays for sensitive cross-clade detection of HIV-1 as reported in the literature and elucidating regions of improved cross-subtype specificity. Inosine and mixed nucleotide bases were included at polymorphic sites. Real-time RT-qPCR and qPCR were performed on plasma viral RNA and cellular lysates. A step-up amplification approach to allow binding of primers across polymorphic regions showed improved sensitivity compared to universal cycling. Unlike a lead competing laboratory-developed assay, all major HIV-1 subtypes, and a wid

SUBMITTER: Kibirige CN 

PROVIDER: S-EPMC8799642 | biostudies-literature | 2022 Jan

REPOSITORIES: biostudies-literature