DETECTION OF HIV-1 BY DIGOXIGENIN-LABELED PCR AND MICROTITRE PLATE SOLUTION HYBRIDIZATION ASSAY AND PREVENTION OF PCR CARRY-OVER BY URACIL-N-GLYCOSYLASE

被引：9

作者：

KING, JA ^{[1
]}

BALL, JK ^{[1
]}

机构：

[1] E BIRMINGHAM DIST GEN HOSP, NATL HLTH SERV TRUST, REG VIRUS LAB, BORDESLEY GREEN E, BIRMINGHAM B9 5ST, W MIDLANDS, ENGLAND

来源：

JOURNAL OF VIROLOGICAL METHODS | 1993年 / 44卷 / 01期

基金：

英国医学研究理事会;

关键词：

HIV-1; PCR; HYBRIDIZATION; UNG; DIGOXIGENIN;

D O I：

10.1016/0166-0934(93)90008-F

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

An extremely sensitive and convenient microtitre plate solution hybridisation assay for the detection of HIV-1 PCR products was developed. The PCR product is labelled by direct incorporation of digoxigenin-dUTP and after denaturation is captured by a microtitre plate coated with a streptavidin-linked biotinylated probe. The PCR/probe hybrids are reacted with an alkaline phosphatase conjugated anti-digoxigenin antibody and detected using an alkaline phosphatase enzyme amplification system. The use of uracil-N-glycosylase and dUTP instead of dTTP in the PCR is used to effectively control carry-over from previous PCR products. The assay can detect single HIV-1 DNA molecules in a background DNA of 0.75 mug.

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收藏

页码：67 / 76

页数：10

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