Improvement of immunodetection of the transcription factor C/EBP homologous protein by western blot

Accumulation of misfolded proteins within the endoplasmic reticulum (ER) induces an unfolded protein response (UPR) that either restores homeostasis or triggers apoptosis in case of adaptation failure. The three activated branches of UPR lead to IRE1-, PERK- and ATF6- dependent transcriptional induc...

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Bibliographic Details
Published in:Analytical biochemistry 2020-07, Vol.601, p.113775-113775, Article 113775
Main Authors: Turpin, Jonathan, Frumence, Etienne, El Safadi, Daed, Meilhac, Olivier, Krejbich-Trotot, Pascale, Viranaïcken, Wildriss
Format: Article
Language:English
Subjects:
A549 Cells
Blotting, Western
CHOP
Endoplasmic Reticulum
Endoplasmic Reticulum - chemistry
Humans
Immunodetection
Life Sciences
Microbiology and Parasitology
Nitrocellulose
Transcription Factor CHOP
Transcription Factor CHOP - analysis
Unfolded Protein Response
Western blot
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Summary:Accumulation of misfolded proteins within the endoplasmic reticulum (ER) induces an unfolded protein response (UPR) that either restores homeostasis or triggers apoptosis in case of adaptation failure. The three activated branches of UPR lead to IRE1-, PERK- and ATF6- dependent transcriptional induction of the gene encoding the transcription factor C/EBP homologous protein (CHOP) which plays an important role in apoptosis induction. In conventional immunoblotting conditions, detection of CHOP is a difficult task. Using a fixation step, we have optimized the detection of CHOP and this method provides a valuable tool to decipher CHOP involvement in UPR. [Display omitted] •Low retention of CHOP protein on nitrocellulose membrane during western blot.•Glutaraldehyde (GA) fixation increase reactivity to CHOP in western blot.•GA fixation decrease background.•GA fixation decrease non specific bands.
ISSN:0003-2697
1096-0309
DOI:10.1016/j.ab.2020.113775