title: A resting cell assay for improved detection of antibody-mediated neutralization of HIV type 1 primary isolates.
abstract: The sensitivity with which antibody-mediated neutralization is detected in vitro is dependent on the virus, the antibody, the target cells, and the culture conditions used in the assay. Using activated and transformed target cells, the ability of various culture-adapted and primary strains of HIV-1 to be neutralized by different polyclonal and monoclonal antibody preparations has been thoroughly studied. However, the vast majority of HIV-1-susceptible CD4+ cells in vivo are not activated or transformed, but are quiescent. Because resting lymphocytes can be infected with HIV-1, we initiated studies to determine (1) if the use of resting lymphocytes as target cells would result in a neutralization assay with increased sensitivity, (2) if the degree of target cell activation had a measurable effect on the sensitivity with which antibody-mediated neutralization could be detected, and (3) whether, using a more sensitive assay, neutralizing antibodies in patients' sera might be detectable that had been below the threshold of detection when using "conventional" assays. The experiments described in the studies below reveal that an inverse relationship exists between the level of target cell activation and the sensitivity with which neutralization can be detected. Moreover, using an assay in which unstimulated peripheral blood mononuclear cells serve as target cells, experiments show that antibody-mediated neutralization of primary and prototype laboratory isolates of HIV-1 can be detected with 10- to 100-fold greater sensitivity than when stimulated peripheral blood mononuclear cells are used as target cells. With this resting cell assay, neutralizing activity can be detected in the sera of HIV-positive subjects that, by previously used "conventional" neutralization assays, was undetectable.
authors.affiliation: Veterans Affairs Medical Center, New York, New York 10010, USA.
authors.forename: S
authors.initials: S
authors.lastname: Zolla-Pazner
authors.forename: S
authors.initials: S
authors.lastname: Sharpe
mesh_terms: D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D015483:HIV Antibodies; D015497:HIV-1 / Q000254:growth & development / Q000276:immunology* / Q000302:isolation & purification; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000276:immunology* / Q000382:microbiology; D008213:Lymphocyte Activation; D008969:Molecular Sequence Data; D009500:Neutralization Tests / Q000379:methods*; D012680:Sensitivity and Specificity; D013997:Time Factors
chemical_list: D000911:Antibodies, Monoclonal; D015483:HIV Antibodies