A cell-free assay to determine the stoichiometry of plasma membrane proteins

BioTechniques

Juan P Vivar¹  Sebastian Brauchi¹  Carlos B Gonzalez¹  Cesar Trigo¹ 

[1]1Instituto de Fisiología, Facultad de Medicina, Universidad Austral de Chile, Campus Isla Teja, Valdivia, Chile

关键词: LSCM;    TIRFM;    Stochastic-GFP photobleaching;    membrane proteins;    membrane sheet;    Hv1;   

DOI  :  10.2144/000113977

来源: DOAJ

【 摘 要 】

Plasma membrane receptors, transporters, and ion channel molecules are often found as oligomeric structures that participate in signaling cascades essential for cell survival. Different states of protein oligomerization may play a role in functional control and allosteric regulation. Stochastic GFP-photobleaching (SGP) has emerged as an affordable and simple method to determine the stoichiometry of proteins at the plasma membrane. This non-invasive optical approach can be useful for total internal reflection of fluorescence microscopy (TIRFM), where signal-to-noise ratio is very high at the plasma membrane. Here, we report an alternative methodology implemented on a standard laser scanning confocal microscope (LSCM). The simplicity of our method will allow for its implementation in any epifluorescence microscope of choice.

【 授权许可】

Unknown