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Poly A-linked non-isotopic microtiter plate reverse transcriptase assay for sensitive detection of clinical human immunodeficiency virus isolates.


J Virol Methods. 1995 Nov;55(3):347-56. Unique Identifier : AIDSLINE

A colorimetric reverse transcriptase assay (cRT assay) was developed for quantitative detection of HIV-1. In this format, reverse transcriptase incorporates biotin-labeled dUTP onto oligo-dT primers hybridized to poly A templates. The templates are covalently bound to the surface of microtiter wells. The amount of incorporated biotin-labeled dUTP is measured by binding horseradish peroxidase conjugated streptavidin, washing away unbound peroxidase, adding colorimetric substrate and then reading with a standard colorimetric reader. The sensitivity of the assay is very good. As little as 3 x 10(5) molecules of recombinant HIV-RT can be detected after 20 h of reaction time. Direct comparison using 3 cultured clinical isolates indicates that this level of detection is equivalent to the commercially available p24 antigen capture assay and the HIV-RNA assay based on branched DNA signal amplification. Other retroviruses, such as HIV-2 and feline immunodeficiency virus (FIV), can also be detected in this format. This non-isotopic assay is easy to perform and could provide a convenient and quantitative method for HIV study by monitoring reverse transcriptase, an essential activity in the infection process.

Animal Antigens, Viral/ANALYSIS Colorimetry Comparative Study Evaluation Studies Human HIV/ENZYMOLOGY/ISOLATION & PURIF HIV Core Protein p24/ANALYSIS HIV-1/ENZYMOLOGY/*ISOLATION & PURIF HIV-2/ENZYMOLOGY/*ISOLATION & PURIF Immunodeficiency Virus, Feline/ENZYMOLOGY/ISOLATION & PURIF *Immunoenzyme Techniques Poly A Reagent Kits, Diagnostic RNA-Directed DNA Polymerase/*ANALYSIS RNA, Viral/ANALYSIS Sensitivity and Specificity Time Factors JOURNAL ARTICLE


Information in this article was accurate in August 30, 1996. The state of the art may have changed since the publication date. This material is designed to support, not replace, the relationship that exists between you and your doctor. Always discuss treatment options with a doctor who specializes in treating HIV.