A bioluminescence assay for gene expression by continuously growing mammalian cells: application for detection of human immunodeficiency virus type 1 (HIV-1).
An in-situ assay for monitoring regulated gene expression in continuously growing mammalian cells is described. This technique can be used for the detection of the transactivator (Tat) protein in human immunodeficiency virus(HIV)-infected cells. Human kidney cells 293, harboring the luc gene, and fused to the HIV-1 long terminal repeat, were isolated and served as tester cells. Tat is supplied by transfection with a tat-carrying plasmid, or alternatively by addition of Tat-containing cell extracts, made from virus-infected or plasmid-transfected cells. Light emitted from the tester cells is recorded on film continuously, or by a photo sensor. Transactivation by HIV Tat results in a pronounced increase in light emission from the tester cells (up to 3000-fold). This assay, which detects HIV-specific gene products, may be used as a diagnostic tool for the detection of active HIV present in peripheral blood.
Molecular Genetics Department, Hebrew University-Hadassah Medical School, Jerusalem, Israel.
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Israel
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Honigman
D000818:Animals; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D015870:Gene Expression; D015696:Gene Products, tat / Q000235:genetics; D016325:HIV Long Terminal Repeat / Q000235:genetics; D015497:HIV-1 / Q000235:genetics / Q000302:isolation & purification*; D006801:Humans; D008027:Light; D008156:Luciferases / Q000235:genetics; D008322:Mammals; D010957:Plasmids; D013601:T-Lymphocytes / Q000382:microbiology; D015533:Transcriptional Activation; D054322:tat Gene Products, Human Immunodeficiency Virus
D015696:Gene Products, tat; D054322:tat Gene Products, Human Immunodeficiency Virus; D008156:Luciferases; D015500:Chloramphenicol O-Acetyltransferase