Abstract
Condensin complexes compact and disentangle chromosomes in preparation for cell division. Commercially available antibodies raised against condensin subunits have been widely used to characterise their cellular interactome. Here we have assessed the specificity of a polyclonal antibody (Bethyl A302-276A) that is commonly used as a probe for NCAPH2, the kleisin subunit of condensin II, in mammalian cells. We find that, in addition to its intended target, this antibody cross-reacts with one or more components of the SWI/SNF family of chromatin remodelling complexes in an NCAPH2-independent manner. This cross-reactivity, with an abundant chromatin-associated factor, is likely to affect the interpretation of protein and chromatin immunoprecipitation experiments that make use of this antibody probe.
Highlights
Antibody probes are critical tools for biomedical research
In order to identify proteins that physically interact with the condensin II complex, we performed affinity purification mass spectrometry using a polyclonal antibody raised against the C-terminus of the NCAPH2 subunit (Bethyl Labs A302-276A, referred to hereafter as B276)
Immunoprecipitation was performed from mouse embryonic stem cell whole cell extract, without crosslinking, using either B276 or a rabbit IgG control
Summary
Antibody probes are critical tools for biomedical research. their utility depends on both affinity and specificity for the intended target. We perform immunoprecipitation mass spectrometry followed by control experiments, to show that a commercial antibody marketed for the detection of the condensin II subunit NCAPH2 cross-reacts with a subunit of SWI/SNF.
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