一种用于人类免疫缺陷病毒的高灵敏度化学发光逆转录酶检测法。

A highly sensitive chemiluminescent reverse transcriptase assay for human immunodeficiency virus.

作者信息

Odawara Fumitomo, Abe Hideki, Kohno Takehiro, Nagai-Fujii Youko, Arai Kenji, Imamura Shigeyuki, Misaki Hideo, Azuma Hiroshi, Ikebuchi Kenji, Ikeda Hisami, Mohan Sharad, Sano Kouichi

机构信息

Fine Chemicals and Diagnostic Division, Asahi Kasei Corporation, 632-1 Mifuku, Ohito-cho, Tagata-gun, Shizuoka 410-2321, Japan.

出版信息

J Virol Methods. 2002 Oct;106(1):115-24. doi: 10.1016/s0166-0934(02)00142-8.

DOI:10.1016/s0166-0934(02)00142-8

PMID:12367736

Abstract

A simple and highly sensitive reverse transcriptase (RT) assay was developed by combining a previously reported non-radioisotopic RT assay with the use of a template-primer-immobilized microplate, an enzyme capture protocol, product digestion and a chemiluminescent substrate. The assay was able to detect directly the RT activity in serum samples, plasma and cell culture medium without the need for concentration and extraction of the enzyme. The assay was able to detect RT activity equivalent to 100 virions/ml of HIV-1. These results suggest that this highly sensitive chemiluminescent RT assay can be used not only for virological investigation but also for routine screening of biopharmaceuticals.

摘要

通过将先前报道的非放射性逆转录酶（RT）检测方法与使用固定有模板引物的微孔板、酶捕获方案、产物消化和化学发光底物相结合，开发了一种简单且高度灵敏的RT检测方法。该检测方法能够直接检测血清样本、血浆和细胞培养基中的RT活性，无需对酶进行浓缩和提取。该检测方法能够检测到相当于每毫升100个HIV-1病毒颗粒的RT活性。这些结果表明，这种高度灵敏的化学发光RT检测方法不仅可用于病毒学研究，还可用于生物制药的常规筛选。