Yang J, Thomason D B
Department of Physiology and Biophysics, University of Tennessee Health Science Center, Memphis 38163.
Biotechniques. 1993 Nov;15(5):848-50.
Many reporter gene assays require killing the cell by fixation or lysis. For assays in living cells, the substrate delivery is inefficient and cannot be supplied in situ in a bolus, which makes assays highly variable. We report a simple synthesis of a luciferin ester that is both photolyzable and cleaved by endogenous esterases such that luciferase activity in living cells is easily monitored. Although the photolyzed substrate can be delivered in bolus, the rapid equilibration of the luciferin ester in the cell and the continuous delivery by the endogenous esterases allow stable, long-term measurements of luciferase activity.
许多报告基因检测需要通过固定或裂解来杀死细胞。对于活细胞检测，底物递送效率低下且不能一次性原位供应，这使得检测结果高度可变。我们报告了一种荧光素酯的简单合成方法，该荧光素酯既可光解又可被内源性酯酶裂解，从而能够轻松监测活细胞中的荧光素酶活性。尽管光解后的底物可以一次性递送，但荧光素酯在细胞中的快速平衡以及内源性酯酶的持续递送使得能够对荧光素酶活性进行稳定、长期的测量。
