Ma Duo H, Alto Neal M, Radhakrishnan Arun
Departments of Molecular Genetics and Microbiology, University of Texas Southwestern Medical Center, Dallas, TX, USA.
Department of Microbiology, University of Texas Southwestern Medical Center, Dallas, TX, USA.
Methods Mol Biol. 2025;2888:83-99. doi: 10.1007/978-1-0716-4318-1_7.
Cholesterol is a critical lipid that is present at high concentrations in the plasma membranes of animal cells. Most of the membrane cholesterol is sequestered by other membrane lipids and the transmembrane domains of proteins. Cholesterol in excess of such sequestration forms a pool that is referred to as "accessible cholesterol." This pool of cholesterol plays a crucial role in maintaining lipid homeostasis and in controlling cell growth. The accessible cholesterol pool can also be exploited by bacteria and viruses to promote infection and host immune responses rapidly lower levels of this pool to confer protection. We had previously developed a bacterial toxin sensor called ALOD4 to monitor and quantify accessible cholesterol in cultured cells. Here, we report the characterization of a modified version of ALOD4 that is specialized to detect and monitor accessible cholesterol levels in primary immune cells by flow cytometry analysis.
胆固醇是一种关键脂质，在动物细胞质膜中以高浓度存在。大多数膜胆固醇被其他膜脂质和蛋白质的跨膜结构域隔离。超过这种隔离量的胆固醇形成一个被称为“可及胆固醇”的池。这个胆固醇池在维持脂质稳态和控制细胞生长中起关键作用。细菌和病毒也可以利用这个可及胆固醇池来促进感染，而宿主免疫反应会迅速降低这个池的水平以提供保护。我们之前开发了一种名为ALOD4的细菌毒素传感器，用于监测和量化培养细胞中的可及胆固醇。在此，我们报告了一种经过修饰的ALOD4的特性，该修饰版本专门用于通过流式细胞术分析检测和监测原代免疫细胞中的可及胆固醇水平。
