A novel dual luciferase assay for the simultaneous monitoring of HIV infection and cell viability

作者：Mitsuki Yu ya; Yamamoto Takuya; Mizukoshi Fuminori; Momota Masatoshi; Terahara Kazutaka; Yoshimura Kazuhisa; Harada Shigeyoshi^*; Tsunetsugu Yokota Yasuko^*

来源：Journal of Virological Methods, 2016, 231: 25-33.

DOI：10.1016/j.jviromet.2016.02.006

• HIV-1
• Reporter virus
• Reporter cell
• Renilla luciferase
• Firefly luciferase

Human immunodeficiency virus type 1 (HIV-1) reporter cell lines are critical tools for drug development. However, one disadvantage of HIV-1 reporter cell lines is that reductions in reporter gene activity need to be normalized to cytotoxicity, i.e., live cell numbers. Here, we developed a dual luciferase assay based on a R. reniformis luciferase (hRLuc)-expressing R5-type HIV-1 (NLAD8-hRLuc) and a CEM cell line expressing CCR5 and firefly luciferase (R5CEM-FiLuc). The NLAD8-hRLuc reporter virus was replication competent in peripheral blood mononuclear cells. The level of hRLuc was correlated with p24 antigen levels (p < 0.001, R = 0.862). The target cell line, R5CEM-FiLuc, stably expressed the firefly luciferase (FiLuc) reporter gene and allowed the simultaneous monitoring of compound cytotoxicity. The dual reporter assay combining a NLAD8-hRLuc virus with R5CEM-FiLuc cells permitted the accurate determination of drug susceptibility for entry, reverse transcriptase, integrase, and protease inhibitors at different multiplicities of infection. This dual reporter assay provides a rapid and direct method for the simultaneous monitoring of HIV infection and cell viability.

• 出版日期2016-5