title: Development of a safe and rapid neutralization assay using murine leukemia virus pseudotyped with HIV type 1 envelope glycoprotein lacking the cytoplasmic domain.
abstract: Neutralizing antibody (NAb) is a critical component of an immune system that can potentially provide sterilizing protection against human immunodeficiency virus type 1 (HIV-1). Therefore, an in vitro assay that can rapidly, safely, and accurately evaluate the NAb response vaccine candidates elicit, especially against a large number of HIV-1 variants, would be highly valuable. It has been demonstrated that HIV-1 envelope glycoprotein lacking the cytoplasmic domain can pseudotype murine leukemia virus encoding the beta-galactosidase gene and that this pseudovirus can specifically infect CD4(+) cells (Schnierle BS, Stitz J, Bosch V, et al.: Proc Natl Acad Sci USA 1997;94:8640-8645). Because the pseudovirus is not biohazardous and because the infection can be quantitatively determined within 2 days, we examined the feasibility of using the pseudovirus for high-throughput neutralization assays for HIV-1. We have generated viruses pseudotyped with gp140 of six different HIV-1 isolates (LAI, RF, Bal, AD8, 89.6, and DH12). All six pseudoviruses were infectious and exhibited expected coreceptor usage phenotype in HOS-CD4 cells expressing either CCR5 or CXCR4. More importantly, the neutralization sensitivity profile of these pseudoviruses was virtually identical to that observed from more conventional neutralization assays using either HIV-1 or SHIV. All pseudoviruses could be neutralized by broadly reactive human monoclonal antibody IgG1 b12. Our results indicate that the pseudoviruses are ideal for high-throughput evaluation of immune sera for their capacity to broadly neutralize a large number of HIV-1 isolates.
authors.affiliation: Laboratory of Molecular Microbiology, NIAID, NIH, Bethesda, Maryland 20892, USA.
authors.forename: Y B
authors.initials: YB
authors.lastname: Kim
authors.forename: M K
authors.initials: MK
authors.lastname: Lee
authors.forename: D P
authors.initials: DP
authors.lastname: Han
authors.forename: M W
authors.initials: MW
authors.lastname: Cho
mesh_terms: D001483:Base Sequence; D015153:Blotting, Western; D003593:Cytoplasm / Q000276:immunology*; D017931:DNA Primers; D015699:HIV Envelope Protein gp120 / Q000276:immunology*; D006801:Humans; D009052:Leukemia Virus, Murine / Q000276:immunology*; D009500:Neutralization Tests
chemical_list: D017931:DNA Primers; D015699:HIV Envelope Protein gp120