PDF(Pubmed)
Abstract：
BAX activation techniques are crucial to studying the intrinsic pathway of apoptosis- thousands of pro-apoptotic signals converge on BAX activation. Current methodologies are predominantly limited to membrane permeabilization studies, which assess endpoint functionality of oligomeric BAX, but overlook early activation steps of cytosolic BAX. Here we detail FLAMBE: a fluorescence polarization ligand assay for monitoring BAX early-activation in solution. We also describe a dual-metric parameterization strategy for distillation of kinetic data and comparative analyses when studying candidate ligands. For complete details on the use and execution of this protocol, please refer to Gelles et al. (2022).
摘要：
BAX激活技术对于研究凋亡的内在途径至关重要-成千上万的促凋亡信号集中在BAX激活上。目前的方法主要限于膜透化研究，评估寡聚BAX的终点功能，但忽略了细胞溶质BAX的早期激活步骤。在这里，我们详细介绍了FLAMBE：一种用于监测溶液中BAX早期活化的荧光偏振配体测定。我们还描述了在研究候选配体时用于蒸馏动力学数据和比较分析的双度量参数化策略。有关此协议的使用和执行的完整详细信息，请参考Gelles等人。（2022年）。
