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Updated: Oct 6, 2025
Brain Membrane Fractionation: An Ex Vivo Approach to Assess Subsynaptic Protein Localization
Published on: May 12, 2017
GST-Perfringolysin O production for the localization and quantification of membrane cholesterol in human and mouse
Leire Goicoechea1,2,3, Fabian Arenas1,2, Fernanda Castro1,2
1Department of Cell Death and Proliferation, Institute of Biomedical Research of Barcelona (IIBB), CSIC, Barcelona, Spain.
Abstract:
Abnormal cholesterol metabolism is linked to many neurodegenerative disorders. Here, we present a protocol for the production of a recombinant protein consisting of a Glutathione-S-Transferase tag fused with the Perfringolysin O (PFO). The GST-PFO tag enables analysis of the localization of cholesterol in subcellular membranes of human and mice brain and liver tissues. We have used this approach for samples from Niemann-Pick type C disease and non-alcoholic steatohepatitis models. The construct may also have applications for the diagnosis of cholesterol-accumulating disorders. For complete details on the use and execution of this protocol, please refer to Kwiatkowska et al. (2014).
