title: A novel assay for antibody-dependent cell-mediated cytotoxicity against HIV-1- or SIV-infected cells reveals incomplete overlap with antibodies measured by neutralization and binding assays.
abstract: The resistance of human immunodeficiency virus type 1 (HIV-1) to antibody-mediated immunity often prevents the detection of antibodies that neutralize primary isolates of HIV-1. However, conventional assays for antibody functions other than neutralization are suboptimal. Current methods for measuring the killing of virus-infected cells by antibody-dependent cell-mediated cytotoxicity (ADCC) are limited by the number of natural killer (NK) cells obtainable from individual donors, donor-to-donor variation, and the use of nonphysiological targets. We therefore developed an ADCC assay based on NK cell lines that express human or macaque CD16 and a CD4(+) T-cell line that expresses luciferase from a Tat-inducible promoter upon HIV-1 or simian immunodeficiency virus (SIV) infection. NK cells and virus-infected targets are mixed in the presence of serial plasma dilutions, and ADCC is measured as the dose-dependent loss of luciferase activity. Using this approach, ADCC titers were measured in plasma samples from HIV-infected human donors and SIV-infected macaques. For the same plasma samples paired with the same test viruses, this assay was approximately 2 orders of magnitude more sensitive than optimized assays for neutralizing antibodies-frequently allowing the measurement of ADCC in the absence of detectable neutralization. Although ADCC correlated with other measures of Env-specific antibodies, neutralizing and gp120 binding titers did not consistently predict ADCC activity. Hence, this assay affords a sensitive method for measuring antibodies capable of directing ADCC against HIV- or SIV-infected cells expressing native conformations of the viral envelope glycoprotein and reveals incomplete overlap of the antibodies that direct ADCC and those measured in neutralization and binding assays.
authors.affiliation: Department of Microbiology and Immunobiology, Harvard Medical School, New England Primate Research Center, Southborough, Massachusetts, USA.
authors.forename: Michael D
authors.initials: MD
authors.lastname: Alpert
authors.forename: Lisa N
authors.initials: LN
authors.lastname: Heyer
authors.forename: David E J
authors.initials: DE
authors.lastname: Williams
authors.forename: Jackson D
authors.initials: JD
authors.lastname: Harvey
authors.forename: Thomas
authors.initials: T
authors.lastname: Greenough
authors.forename: Maria
authors.initials: M
authors.lastname: Allhorn
authors.forename: David T
authors.initials: DT
authors.lastname: Evans
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000737:chemistry*; D000920:Antibody-Dependent Cell Cytotoxicity; D015496:CD4-Positive T-Lymphocytes / Q000821:virology; D002460:Cell Line; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D015699:HIV Envelope Protein gp120 / Q000378:metabolism; D015497:HIV-1 / Q000378:metabolism*; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry; D007694:Killer Cells, Natural / Q000821:virology; D008251:Macaca; D009500:Neutralization Tests / Q000379:methods*; D011485:Protein Binding; D017452:Receptors, IgG / Q000096:biosynthesis; D015203:Reproducibility of Results; D015302:Simian Immunodeficiency Virus / Q000378:metabolism*; D014773:Virology / Q000379:methods
chemical_list: D000914:Antibodies, Viral; D015699:HIV Envelope Protein gp120; D007074:Immunoglobulin G; D017452:Receptors, IgG