PMID: 8609200Nov 1, 1995Paper

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

Journal of Virological Methods
K SuzukiM Imai

Abstract

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.

References

Dec 1, 1992·Journal of Virological Methods·J Eberle, R Seibl
Aug 1, 1991·Proceedings of the National Academy of Sciences of the United States of America·M E GoldmanJ M Hoffman
Feb 1, 1991·Journal of Virological Methods·T PorstmannG Sydow
Oct 1, 1993·Journal of Virological Methods·K SuzukiR T Steigbigel
Jul 1, 1994·Kansenshōgaku zasshi. The Journal of the Japanese Association for Infectious Diseases·T NakanoM Nakai

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Citations

Oct 9, 2002·Journal of Virological Methods·Fumitomo OdawaraKouichi Sano
Dec 16, 2006·Retrovirology·Nicole M WillemsenDavid Harrich
Oct 30, 2016·Molecular Therapy. Methods & Clinical Development·Samantha A McAlleryStuart G Turville
May 15, 1999·Journal of Clinical Microbiology·J F SearsA S Khan

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