Jarvela Timothy S, Linstedt Adam D
Department of Biological Sciences, Carnegie Mellon University, Pittsburgh, Pennsylvania.
Curr Protoc Cytom. 2014 Jul 1;69:12.35.1-12.35.10. doi: 10.1002/0471142956.cy1235s69.
Generating loss of protein function is a powerful investigatory tool particularly if carried out on a physiologically relevant timescale in a live-cell fluorescent imaging experiment. KillerRed mediated chromophore assisted light inactivation (CALI) uses genetic encoding for specificity and light for acute inactivation that can also be spatially restricted. This unit provides protocols for setting up and carrying out properly controlled KillerRed experiments during live-cell imaging of cultured cells.
产生蛋白质功能丧失是一种强大的研究工具，特别是在活细胞荧光成像实验中，在生理相关的时间尺度上进行时。KillerRed介导的发色团辅助光灭活（CALI）利用基因编码实现特异性，并利用光进行急性灭活，这种灭活还可以在空间上受到限制。本单元提供了在培养细胞的活细胞成像过程中设置和进行适当控制的KillerRed实验的方案。
