High-Throughput Quantitation of Intracellular Trafficking and Organelle Disruption by Flow Cytometry
Pei Zhi Cheryl Chia, Yasmin M Ramdzan, Fiona J Houghton, Danny M Hatters, Paul A Gleeson
TRAFFIC | WILEY | Published : 2014
Current methods for the quantitation of membrane protein trafficking rely heavily on microscopy, which has limited quantitative capacity for analyses of cell populations and is cumbersome to perform. Here we describe a simple flow cytometry-based method that circumvents these limitations. The method utilizes fluorescent pulse-width measurements as a highly sensitive indicator to monitor the changes in intracellular distributions of a fluorescently labelled molecule in a cell. Pulse-width analysis enabled us to discriminate cells with target proteins in different intracellular locations including Golgi, lyso-endosomal network and the plasma membrane, as well as detecting morphological changes..View full abstract
We thank Dorothee Bourges for expert advice. This work was supported by funding from the Australian Research Council and National Health and Medical Research Council of Australia. P.Z.C.C. was supported by a Melbourne International Graduate Scholarship.