Development of a method for correlating the presence of HIV antigen and provirus.

Important note: Information in this article was accurate in 1990. The state of the art may have changed since the publication date.

Development of a method for correlating the presence of HIV antigen and provirus.

Int Conf AIDS. 1989 Jun 4-9;5:434 (abstract no. Th.B.P.111). Unique Identifier : AIDSLINE ICA5/00221989
Paul DB; Bankowski M; Kuhns M; McNamara A; Landay AL; Spear GT; Rush University, Chicago, IL, USA

Abstract: OBJECTIVE: a) To develop a sensitive flow cytometric assay that will allow detection of HIV antigen containing cells. b) To correlate the presence of HIV antigen with provirus in cells. METHODS: Cell lines were fixed, permeabilized, and stained for HIV antigens with a variety of fluorescent antibodies. After staining, fluorescence was quantified by flow cytometry. In some experiments, populations of infected and uninfected cells were mixed and stained, and the positive and negative populations identified using the flow cytometer. The detection of provirus in fixed cells was accomplished by DNA amplification, using the polymerase chain reaction (PCR) and a liquid phase molecular hybridization system. RESULTS: We compared 3 antibody systems: biotinylated lgG from AIDS patients; affinity purified rabbit anti-p24, and a monoclonal antibody against p24, each with an appropriate secondary reagent. All 3 systems enabled detection of HIV infected cells. The monoclonal antibody had the greatest sensitivity for detection of infection in cells, as shown by increased fluorescence over control antibodies or uninfected cells. Provirus was detectable by PCR in fixed and stained cells. This technique is appropriate for use in flow cytometric separation of cells into positive and negative populations and will allow analysis for provirus by PCR. CONCLUSIONS: The anti-p24 monoclonal antibody allowed highly sensitive detection of cells containing p24. Analysis by PCR, followed by liquid phase molecular hybridization, confirmed the HIV provirus was detectable after fixation. This method will prove useful in determining the correlation between the presence of p24 and the provirus in cells from HIV infected individuals.

Keywords: Antibodies, Monoclonal/DIAGNOSTIC USE Flow Cytometry Gene Products, gag/ANALYSIS Human HIV Antigens/*ANALYSIS HIV Infections/*IMMUNOLOGY/MICROBIOLOGY Polymerase Chain Reaction Proviruses/*ISOLATION & PURIF Viral Core Proteins/ANALYSIS ABSTRACT
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