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A Guide to Modern Quantitative Fluorescent Western Blotting with Troubleshooting Strategies
Published on: November 20, 2014
A guide to selecting high-performing antibodies for TMEM175 (UniProt ID: Q9BSA9) for use in western blot,
Vera Ruíz Moleón1, Charles Alende1, Maryam Fothouhi1
1Department of Neurology and Neurosurgery, Structural Genomics Consortium, The Montreal Neurological Institute, McGill University, Montreal, Quebec, Canada.
Abstract:
TMEM175 is the pore-forming subunit of a lysosomal K+ channel complex that regulates lysosomal pH stability and membrane potential. To further investigate its cellular functions and implications in neurodegenerative diseases, antibody reagents are needed. Here we have characterized six TMEM175 commercial antibodies for western blot, immunoprecipitation, and immunofluorescence using a standardized experimental protocol based on comparing read-outs in knockout cell lines and isogenic parental controls. These studies are part of a larger, collaborative initiative seeking to address antibody reproducibility issues by characterizing commercially available antibodies for human proteins and publishing the results openly as a resource for the scientific community. While use of antibodies and protocols vary between laboratories, we encourage readers to use this report as a guide to select the most appropriate antibodies for their specific needs.

