title: Polymerase chain reaction-based assay for antibody-mediated neutralization of HIV-1 reveals a population of nonneutralized virus undetected by conventional p24 assay.
abstract: To be successful with strategies involving passive immunization or the generation of neutralizing antibodies against HIV, it is crucial that we improve our understanding of the process of antibody-mediated HIV neutralization. We have used a neutralization assay based on polymerase chain reaction (PCR) that is more rapid and sensitive than the conventional p24 neutralization assay based on enzyme-linked immunosorbent assay (ELISA). PCR assays permit measurement of the number of infectious events and can detect small amounts of HIV-1 only a few days postinfection. In these studies, the human anti-V3 monoclonal antibody 694/98-D was used to neutralize the infectivity of the laboratory isolate HIVIIIB for CEM-SS cells. 8E5/LAV cells, which contain a single integrated copy of proviral DNA per cell, served as a standard to determine the amount of HIV-1 copies in infected CEM-SS cells. Evaluation of antibody-mediated neutralization was possible at 2 to 3 days postinfection, at a time when p24 readouts were not conclusive. We achieved >95% neutralization of HIVIIIB, and of its molecular clone HXB2, using the monoclonal antibody 694/98-D. This degree of neutralization is probably highly significant in vivo. Nevertheless, a small amount of both HIVIIIB and HXB2 ( approximately 5%) escapes neutralization and can consistently be detected after a few days by this sensitive assay. Experiments with different anti-HIV monoclonal antibodies and viruses showed that the assay could be applied to anti-V3 as well as anti-CD4 binding domain antibodies as well as HIV laboratory strains or primary isolates.
authors.affiliation: New York Veterans Affairs Medical Center and Departments of Medicine and Pathology, New York University, New York City, New York 10010, USA.
authors.forename: J M
authors.initials: JM
authors.lastname: Achkar
authors.forename: X H
authors.initials: XH
authors.lastname: Wang
authors.forename: P
authors.initials: P
authors.lastname: Nyambi
authors.forename: M K
authors.initials: MK
authors.lastname: Gorny
authors.forename: S
authors.initials: S
authors.lastname: Zolla-Pazner
authors.forename: J C
authors.initials: JC
authors.lastname: Bandrés
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000494:pharmacology; D000956:Antigens, Viral / Q000276:immunology; D015139:Blotting, Southern; D004279:DNA, Viral / Q000032:analysis; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D015483:HIV Antibodies / Q000276:immunology* / Q000494:pharmacology; D016655:HIV Core Protein p24 / Q000032:analysis; D015699:HIV Envelope Protein gp120 / Q000276:immunology; D015497:HIV-1 / Q000187:drug effects / Q000276:immunology* / Q000302:isolation & purification*; D006801:Humans; D019169:Jurkat Cells; D009500:Neutralization Tests; D010446:Peptide Fragments / Q000276:immunology; D016133:Polymerase Chain Reaction / Q000379:methods*; D011533:Proviruses / Q000235:genetics; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D000911:Antibodies, Monoclonal; D000956:Antigens, Viral; D004279:DNA, Viral; D015483:HIV Antibodies; D016655:HIV Core Protein p24; D015699:HIV Envelope Protein gp120; C059762:HIV envelope protein gp120 (305-321); D010446:Peptide Fragments