Ying Guo1, Xiao-wei Wang, Yang Xu, Xiao Lv, Rui-ping Wang, Jun-yi Liu. 1. Department of Chemical Biology, School of Pharmaceutical Science, State Key laboratory of Natural and Biomimetic Drugs, Peking University, Beijing 100083, China.
Abstract
OBJECTIVE: to establish an ELISA method for detecting the activity of reverse transcriptase. METHODS: Oligo (dT) 15 was immobilized via its 5'-terminal phosphate to Covalink-NH microtiter plates,The biotin-dUTP was incorporated by reverse transcriptase. The products were detected and quantified using a colorimetric streptavidin-alkaline phosphatase reporter system. RESULTS: We established an ELISA RT assay method and tested the inhibition activity of PFA and NVP respectively. Using this method we also studied the kinetic mechanism of AMV-RT and HIV-1 RT. At the same time, we evaluated the stability, repeatability, specificity and sensitivity of this ELISA method. CONCLUSION: This ELISA method has particular advantages, such as simplicity, rapidity, specificity and little pollution, and is suitable for screening and studying of specific inhibitors of HIV-1 reverse transcriptase and other RTs.
OBJECTIVE: to establish an ELISA method for detecting the activity of reverse transcriptase. METHODS:Oligo (dT) 15 was immobilized via its 5'-terminal phosphate to Covalink-NH microtiter plates,The biotin-dUTP was incorporated by reverse transcriptase. The products were detected and quantified using a colorimetric streptavidin-alkaline phosphatase reporter system. RESULTS: We established an ELISA RT assay method and tested the inhibition activity of PFA and NVP respectively. Using this method we also studied the kinetic mechanism of AMV-RT and HIV-1 RT. At the same time, we evaluated the stability, repeatability, specificity and sensitivity of this ELISA method. CONCLUSION: This ELISA method has particular advantages, such as simplicity, rapidity, specificity and little pollution, and is suitable for screening and studying of specific inhibitors of HIV-1 reverse transcriptase and other RTs.