Comparative study of commercially available and homemade anti-VAMP7 antibodies using CRISPR/Cas9-depleted HeLa cells and VAMP7 knockout mice

F1000Research
Agathe VerraesSebastien Nola

Abstract

VAMP7 (vesicle-associated membrane protein) belongs to the intracellular membrane fusion SNARE (Soluble N-ethylmaleimide-sensitive factor attachment protein receptors) protein family. In this study, we used CRISPR/Cas9 genome editing technology to generate VAMP7 knockout (KO) human HeLa cells and mouse KO brain extracts in order to test the specificity and the background of a set of commercially available and homemade anti-VAMP7 antibodies. We propose a simple profiling method to analyze western blotting and use visual scoring for immunocytochemistry staining to determine the extent of the antibodies' specificity. Thus, we were able to rank the performance of a set of available antibodies and further showed an optimized procedure for VAMP7 immunoprecipitation, which we validated using wild-type and KO mouse brain extracts.

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Citations

Dec 29, 2020·Cell Reports·José WojnackiThierry Galli
Apr 23, 2021·The Journal of Cell Biology·Shanna L BowmanMichael S Marks

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Software Mentioned

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RGEN Cas designer
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