Expression and purification of C14orf119 and generation of its polyclonal antibody

C14orf119 is a functionally uncharacterized mitochondrial protein potentially implicated in ischemic stroke pathogenesis. To enable comprehensive biological studies, we developed and validated a specific polyclonal antibody against this target. Our approach involved prokaryotic expression and chromatographic purification of both His- and GST-tagged C14orf119 in Escherichia coli. The purified His-C14orf119 served as immunogen for rabbit polyclonal antibody production, while the purified GST-C14orf119 enabled subsequent affinity purification of target-specific antibody. Rigorous characterization demonstrated the antibody's exclusive specificity for both exogenous and endogenous C14orf119. Notably, the C14orf119 antibody failed to detect control His-tagged proteins, confirming effective removal of anti-His antibodies during the GST-based affinity purification process. This rigorously validated antibody provides a critical molecular tool for functional characterization of C14orf119, paving the way for mechanistic investigations into its pathophysiological roles.