Titration of human immunodeficiency virus type 1 (HIV-1) and quantitative analysis of virus expression in vitro using liquid RNA-RNA hybridization.
An assay is described for titration of human immunodeficiency virus type 1 (HIV-1) and for quantitative analysis of virus expression in vitro. The assay utilizes a liquid RNA-RNA hybridization method coupled with reversible target capture (RTC) on oligo(dT) derivatized magnetic particles. The assay provides a rapid, specific, and sensitive method for quantitation of HIV-1 RNA molecules present either in cells or in viral particles from cell-free culture media. Chronically infected monocytoid U1.1 cells were found to carry 52 pg HIV-1 RNA per 200,000 cells (160 HIV-1 RNA molecules per cell). In contrast, acutely infected CEM and H9 cells carried 3010 and 4370 pg HIV-1 RNA per 200,000 cells (9040 and 13,110 HIV-1 RNA molecules per cell, respectively). No hybridization was observed with uninfected cells or cells infected with HIV-2, HTLV-I, HTLV-II, or EBV. Use of liquid HIV-1 RNA hybridization in association with HIV-1 protein detection methods permits more complete characterization of HIV-1 expression in host cells than either method alone, and also provides a method for standardizing preparations of virus.
Molecular Virology Laboratory, St. Luke's/Roosevelt Hospital Center, Columbia University, New York, New York.
D J
DJ
Volsky
M G
MG
Pellegrino
G
G
Li
K A
KA
Logan
J E
JE
Aswell
N P
NP
Lawrence
S R
SR
Decker
D000163:Acquired Immunodeficiency Syndrome / Q000235:genetics; D015870:Gene Expression; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D009693:Nucleic Acid Hybridization; D012367:RNA, Viral / Q000378:metabolism*; D012680:Sensitivity and Specificity; D012997:Solvents; D014407:Tumor Cells, Cultured
D012367:RNA, Viral; D012997:Solvents