title: Neutralization sensitivity of cell culture-passaged simian immunodeficiency virus.
abstract: CEMx174- and C8166-45-based cell lines which contain a secreted alkaline phosphatase (SEAP) reporter gene under the control of a tat-responsive promoter derived from either SIVmac239 or HIV-1(NL4-3) were constructed. Basal levels of SEAP activity from these cell lines were low but were greatly stimulated upon transfection of tat expression plasmids. Infection of these cell lines with simian immunodeficiency virus (SIV) or human immunodeficiency virus type 1 (HIV-1) resulted in a dramatic increase in SEAP production within 48 to 72 h that directly correlated with the amount of infecting virus. When combined with chemiluminescent measurement of SEAP activity in the cell-free supernatant, these cells formed the basis of a rapid, sensitive, and quantitative assay for SIV and HIV infectivity and neutralization. Eight of eight primary isolates of HIV-1 that were tested induced readily measurable SEAP activity in this system. While serum neutralization of cloned SIVmac239 was difficult to detect with other assays, neutralization of SIVmac239 was readily detected at low titers with this new assay system. The neutralization sensitivities of two stocks of SIVmac251 with different cell culture passage histories were tested by using sera from SIV-infected monkeys. The primary stock of SIVmac251 had been passaged only twice through primary cultures of rhesus monkey peripheral blood mononuclear cells, while the laboratory-adapted stock had been extensively passaged through the MT4 immortalized T-cell line. The primary stock of SIVmac251 was much more resistant to neutralization by a battery of polyclonal sera from SIV-infected monkeys than was the laboratory-adapted virus. Thus, SIVmac appears to be similar to HIV-1 in that extensive laboratory passage through T-cell lines resulted in a virus that is much more sensitive to serum neutralization.
authors.affiliation: New England Regional Primate Research Center, Harvard Medical School, Southborough, Massachusetts 01772-9102, USA.
authors.forename: R E
authors.initials: RE
authors.lastname: Means
authors.forename: T
authors.initials: T
authors.lastname: Greenough
authors.forename: R C
authors.initials: RC
authors.lastname: Desrosiers
mesh_terms: D000469:Alkaline Phosphatase / Q000096:biosynthesis; D000818:Animals; D000914:Antibodies, Viral; D001483:Base Sequence; D002460:Cell Line; D017931:DNA Primers; D017930:Genes, Reporter; D015756:Genes, tat; D016325:HIV Long Terminal Repeat; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D007700:Kinetics; D008253:Macaca mulatta; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D011401:Promoter Regions, Genetic; D011533:Proviruses / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D012091:Repetitive Sequences, Nucleic Acid; D016097:Simian Acquired Immunodeficiency Syndrome / Q000276:immunology; D015302:Simian Immunodeficiency Virus / Q000235:genetics*; D013997:Time Factors; D014162:Transfection
chemical_list: D000914:Antibodies, Viral; D017931:DNA Primers; D011993:Recombinant Fusion Proteins; D000469:Alkaline Phosphatase