IMMUNOLOGICAL CHARACTERIZATION AND DETECTION OF THE HUMAN IMMUNODEFICIENCY VIRUS CORE PROTEIN p24 WITH MURINE MONOCLONAL ANTIBODIES

Abstract

Sixteen murine monoclonal antibodies (MAb) reactive with HIV-1 viral lysate were generated from a mouse immunized with detergent disrupted viral particles. These immunological reagents were thoroughly characterized and tested as capture antibodies in an ELISA for the detection of HIV-1ɪɪɪʙ P, HIV-2ɴɪʜ-ᴢ p26 and SIVᴀɢᴍ p27 antigens in viral lysates. All of the MAb are capable of capturing p24, and five of the MAb can also capture either p26, p27 or both viral antigens. One of these cross-reactive MAb, 7-D4, can detect HIV-1 viral lysate in the pg/ml range and HIV-2 and Sly lysates in the ng/ml range. Full- and partial -length recombinant p24 peptides derived from a subgenomic library of HIV-1ʀꜰ were also probed with the purified MAb in Western Blot, and the results indicate that these MAb exhibit epitope ecificities spanning most of the p24 molecule. Several MAb recognize epitopes at or near the NH₂- and COOH-termini of the protein, and the remaining antibodies react with internal epitopes. In addition, epitope mapping using sequential synthetic peptides (9- mers) that collectively comprise the entire p24 molecule has revealed that MAb 7-D4 recognizes an epitope that is linear in nature and conserved between the HIV-1, HIV-2 and SIV lysates tested. The region containing this epitope is predicted to form a β-turn secondary protein structure, and the immunoreactivity of the epitope itself is not destroyed by denaturants. In summary, a large variety of epitope specificities are represented by the monoclonal antibodies, and this way aid in the detection, characterization and differentiation of HIV isolates, as well as isolates of other closely related lentiviruses.