Proof of principle: An HIV p24 microsphere immunoassay with potential application to HIV clinical diagnosis

✍ Scribed by Pascale Ondoa; Chris Vereecken; Eugene Lekeawung Asahchop; Amber Litzroth; Aziz Diallo; Katrien Fransen; Tandakha Dieye; Robin Ryder; Souleymane Mboup; Luc Kestens

• Publisher: John Wiley and Sons
• Year: 2009
• Tongue: English
• Weight: 263 KB
• Volume: 76B
• Category: Article
• ISSN: 1552-4949
• DOI: 10.1002/cyto.b.20466

✦ Synopsis

Abstract

The measurement of CD4 counts and viral loads on a single instrument such as an affordable flow cytometer could considerably reduce the cost related to the follow‐up of antiretroviral therapy in resource‐poor settings. The aim of this study was to assess whether the HIV‐1 p24 antigen could be measured using a microsphere‐based flow cytometric (FC) assay and the experimental conditions necessary for processing plasma samples. A commercial anti‐p24 antibody pair from Biomaric was used to develop a p24 microsphere immunoassay (MIA) using HIV culture supernatant as the source of antigen. The ultrasensitive Perkin Elmer enzyme immunoassay (EIA) served as a reference assay. Quantification of HIV p24 using the heat‐mediated immune complex disruption format described for plasma samples was feasible using the Biomaric MIA and applicable to a broad range of HIV‐1 Group M subtypes. The inclusion of a tyramide amplification step was successful and increased the fluorescence signal up to 3 logs as compared with the MIA without amplification. The analytical sensitivity of this ultrasensitive Biomaric assay reached 1 pg/mL, whereas the ultrasensitive Perkin Elmer EIA was sensitive to less than 0.17 pg/mL. Our data indicate, for the first time, that the principle of p24 detection using the heat‐denatured ultrasensitive format can be applied to FC. © 2008 Clinical Cytometry Society