Doysabas Karla Cristine C, Oba Mami, Ishibashi Tomoki, Shibata Hideki, Takemae Hitoshi, Shimoda Hiroshi, Tarigan Ronald, Mizutani Tetsuya, Iida Atsuo, Hondo Eiichi
Graduate School of Bioagricultural Sciences, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, Aichi 464-8601, Japan.
Laboratory of Veterinary Microbiology, Cooperative Department of Veterinary Medicine, Tokyo University of Agriculture and Technology, Sawai, Fuchu, Tokyo 183-8509, Japan.
J Vet Med Sci. 2020 Mar 24;82(3):387-393. doi: 10.1292/jvms.20-0021. Epub 2020 Feb 12.
Adenosine 5'-triphosphate (ATP), the major energy currency of the cell, is involved in many cellular processes, including the viral life cycle, and can be used as an indicator of early signs of cytopathic effect (CPE). In this study, we demonstrated that CPE can be analyzed using an FRET-based ATP probe named ATP indicator based on Epsilon subunit for Analytical Measurements (ATeam). The results revealed that as early as 3 hr, the virus infected cells showed a significantly different Venus/cyan fluorescent protein (CFP) ratio compared to the mock-infected cells. The ATeam technology is therefore useful to determine the early signs of ATP-based CPE as early as 3 hr without morphology-based CPE by light microscopy, and enables high throughput determination of the presence of microorganisms in neglected samples stored in laboratories.
腺苷 5'-三磷酸(ATP)是细胞的主要能量货币,参与许多细胞过程,包括病毒生命周期,并且可用作细胞病变效应(CPE)早期迹象的指标。在本研究中,我们证明可以使用一种基于荧光共振能量转移(FRET)的ATP探针——基于ε亚基的分析测量ATP指示剂(ATeam)来分析CPE。结果显示,早在3小时时,与模拟感染细胞相比,病毒感染细胞的金星/青色荧光蛋白(CFP)比率就有显著差异。因此,ATeam技术可用于早在3小时时确定基于ATP的CPE的早期迹象,而无需通过光学显微镜基于形态学的CPE,并且能够高通量确定实验室中储存的被忽视样本中微生物的存在情况。