Morgan J R, Lee J, Tompkins R G, Yarmush M L
Surgical Services, Massachusetts General Hospital, Boston 02114.
Biotechnol Prog. 1994 Jul-Aug;10(4):441-6. doi: 10.1021/bp00028a014.
A method for measuring the concentration of recombinant retrovirus encoding the Escherichia coli LacZ gene is described. The assay is based on the quantitative measurement of beta-galactosidase activity in extracts of cells infected with the LacZ-encoding retrovirus. LacZ-encoded beta-galactosidase activity in transduced cells is measured using the colorimetric substrate, o-nitrophenyl beta-D-galactopyranoside (ONPG), and the results are read using an ELISA plate-reader. Because the entire assay is performed in a 96-well plate, large numbers of samples are easily measured, and the assay has the advantages of rapidity, precision, and ease of data collection. The assay was used to determine the optimal concentration of polybrene, a polycation known to enhance the infectivity of retroviruses, and can be used to evaluate other factors that affect infection, as well as the optimal conditions for production of the recombinant retrovirus.
本文描述了一种用于测量编码大肠杆菌LacZ基因的重组逆转录病毒浓度的方法。该测定基于对感染了编码LacZ的逆转录病毒的细胞提取物中β-半乳糖苷酶活性的定量测量。使用比色底物邻硝基苯基β-D-吡喃半乳糖苷（ONPG）测量转导细胞中LacZ编码的β-半乳糖苷酶活性，并使用酶联免疫吸附测定（ELISA）酶标仪读取结果。由于整个测定在96孔板中进行，因此可以轻松测量大量样品，并且该测定具有快速、精确和易于数据收集的优点。该测定用于确定聚凝胺（一种已知可增强逆转录病毒感染性的聚阳离子）的最佳浓度，并且可用于评估影响感染的其他因素以及重组逆转录病毒生产的最佳条件。
