Protein extracts were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) on 7.5%–16% gradient gels and transferred by semi-dry blotting to Hybond ECL nitrocellulose membranes (GE Healthcare). The following primary antibodies were used to develop the blots: anti-MK2, anti-p38, anti-pT180/Y182-p38, anti-pT334-MK2, anti-p(S/T)-PKD-substrate, anti-Hsp27, anti-pS82-Hsp27 (all purchased from Cell Signaling), anti-GAPDH (Millipore), anti-eEF2, anti-GFP, anti-hRRP40 (all from Santa Cruz Biotechnology), anti-RBM7, anti-ZCCHC8 (both from Atlas antibodies), anti-pS136-Rbm7 (see above); anti-NOGO-B (Rousseau et al. 2005 (link)), and anti-CBP80 (Andersen et al. 2013 (link)) antibody were described before. The anti-FLAG (M2) antibody was purchased from Agilent. ECL detection was carried out on a chemo-luminescence detector (LAS 3000 FujiFilm).
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Western Blot Protein Analysis
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Other organizations : Medizinische Hochschule Hannover, Aarhus University, University of Dundee, MRC Protein Phosphorylation and Ubiquitylation Unit
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Variable analysis
independent variables
- Protein extracts were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) on 7.5%–16% gradient gels
- Measurement of protein levels and phosphorylation states using specific antibodies: anti-MK2, anti-p38, anti-pT180/Y182-p38, anti-pT334-MK2, anti-p(S/T)-PKD-substrate, anti-Hsp27, anti-pS82-Hsp27, anti-GAPDH, anti-eEF2, anti-GFP, anti-hRRP40, anti-RBM7, anti-ZCCHC8, anti-pS136-Rbm7, anti-NOGO-B, anti-CBP80
- Protein extracts were transferred by semi-dry blotting to Hybond ECL nitrocellulose membranes (GE Healthcare)
- ECL detection was carried out on a chemo-luminescence detector (LAS 3000 FujiFilm)
- Anti-FLAG (M2) antibody
- Not explicitly mentioned
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