Western blot analysis was performed as described previously [48 (link)] using the following antibodies: cIAP1 (R&D Systems, Inc., Wiesbaden, Germany), RIP1 (BD Biosciences, Heidelberg, Germany), RIP3 (Novus Biologicals, Littleton, CO, USA), MLKL (GeneTex, Irvine, CA, USA), phospho-MLKL (Cell Signaling Technologies, Danvers, MA, USA), β-Actin (Sigma-Aldrich) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) (HyTest, Turku, Finland) as loading controls and secondary antibodies conjugated to horseradish peroxidase (Santa Cruz Biotechnology, Santa Cruz, CA, USA). Enhanced chemiluminescence was used for detection (Amersham Bioscience, Freiburg, Germany). Alternatively, secondary antibodies labeled with IRDye infrared dyes were used for fluorescence detection (Odyssey Imaging System, LI-COR Bioscience, Bad Homburg, Germany). All Western blots shown are representative of at least two independent experiments.
Glyceraldehyde 3 phosphate dehydrogenase gapdh
Manufactured by HyTest
Sourced in Finland
Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) is an enzyme that catalyzes the sixth step of glycolysis, the metabolic pathway that converts glucose into energy. GAPDH facilitates the conversion of glyceraldehyde 3-phosphate to 1,3-bisphosphoglycerate.
Automatically generated - may contain errors
Lab products found in correlation
Odyssey imaging system, by LI COR (1 mentions)
Horseradish peroxidase, by Santa Cruz Biotechnology (1 mentions)
Enhanced chemiluminescence, by Cytiva (1 mentions)
β actin, by Merck Group (1 mentions)
Ciap1, by R&D Systems (1 mentions)
Phospho mlkl, by Cell Signaling Technology (1 mentions)
Inhibitor mix m, by Serva Electrophoresis (1 mentions)
2 protocols using glyceraldehyde 3 phosphate dehydrogenase gapdh
1
Western Blot Analysis of Cell Signaling
2
Quantifying Protein Expressions in Frozen Tissues
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Frozen tissue samples were homogenized in radioimmunoprecipitation assay buffer containing a mixture of protease inhibitor (inhibitor mix M, Serva, Heidelberg, Germany) and protein expression that was quantified by western blot using specific antibodies to NAD(P)H oxidase (Abcam, Cambridge, UK), MuRF-1 (generous gift of Dr S. Labeit, University Mannheim, Germany), and MAFbx (generated in rabbits against the following peptide sequence CYPRKEQYGDTLQL, Eurogentec, Seraing, Belgium). After incubation with a horseradish peroxidase-conjugated secondary antibody, specific bands were visualized by enzymatic chemiluminescence (Super Signal West Pico, Pierce, Bonn, Germany) and densitometry quantified by a one-dimensional scan software package (Scanalytics, Rockville, USA). Loading differences were controlled by reprobing the blot with an antibody against Glyceraldehyde-3-phosphate-dehydrogenase (GAPDH) (Hytest, Turku, Finland).
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