Western blotting procedures has been described previously.48 (link) The primary antibodies used were ATF6 (Abcam, Cambridge, UK, Cat# ab122897), spliced XBP1 (Biolegend, London, UK, Cat# 619502), PERK (Cell signalling, Cat# C33E10), GRP78 (Fisher Scientific Ireland Ltd, Cat# PA1-014A), phospho-eIF2α (Cell signalling, Cat# 9721), total eIF2α (Cell signalling, Cat# 9722) and or β-actin (Sigma, Cat# A-5060) overnight at 4 °C. The membrane was washed three times with PBS-0.05% Tween and further incubated in appropriate horseradish peroxidase-conjugated secondary antibody (Fisher Scientific Ireland Ltd) for 90 min. Signals were detected using Western Lightening Plus ECL (PERKin Elmer, Dublin, Ireland).
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Western Blot Analysis of UPR Markers
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Other organizations : Ollscoil na Gaillimhe – University of Galway
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- Primary antibodies used: ATF6, spliced XBP1, PERK, GRP78, phospho-eIF2α, total eIF2α, β-actin
- Signals detected using Western Lightening Plus ECL
- Membrane was washed three times with PBS-0.05% Tween
- Membrane was further incubated in appropriate horseradish peroxidase-conjugated secondary antibody for 90 min
- No positive or negative controls were explicitly mentioned in the provided information.
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