Total cellular protein from human HSCs or LX2 cell line prepared using radioimmunoprecipitation assay (RIPA) buffer containing protease inhibitor cocktail (Sigma) was separated on SDS-PAGE following standard protocols (Amersham BioSciences, Piscataway, NJ). Antibodies used for Western blotting were: MAT2α2 (Novus Biologicals, CO), MATβ (Sigma) PPARγ (Santa Cruz Biotechology, CA), α-SMA (Novus), MEF2A (Origene), Anti-Phosphorylated Proteins (Pan) antibody (Invitrogen), p-ERK1/2 and B-Raf (Cell Signaling, MA), Type I collagen (Novus), DDK tag antibody (Origene), HA tag antibody (Genecopoeia), α-tubulin (Genetex, Irvine, CA) and actin (Sigma). Detection was done by the chemiluminescence ECL system (Amersham BioSciences). Blots were quantified using the Quantity One™ densitometry program (Bio-Rad laboratories, Hercules, CA) and test protein expression was normalized to α-tubulin control according to previously published reports (Olaso et al, 2001 (link)).
Quantity one densitometry program
Manufactured by Bio-Rad
Quantity One™ is a software program for densitometry analysis. It enables users to quantify and analyze bands or spots in 1D and 2D electrophoresis gels and blots.
Automatically generated - may contain errors
Lab products found in correlation
α tubulin, by Bio-Rad (1 mentions)
Collagen type 1, by Novus Biologicals (1 mentions)
Ecl system, by Cytiva (1 mentions)
Protease inhibitor cocktail, by Merck Group (1 mentions)
α sma, by Novus Biologicals (1 mentions)
Pparγ, by Santa Cruz Biotechnology (1 mentions)
α tubulin, by GeneTex (1 mentions)
2 protocols using quantity one densitometry program
1
Quantitative Western Blot Analysis of Cellular Proteins
2
Quantifying ECL Signal Densitometry
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The ECL signal was quantified with Quantity One densitometry program (Bio-Rad). Phosphorylation data were normalized and expressed as the fold change from nonstimulated samples. All data are expressed as the mean ± SD derived from at least three independent experiments. The t-test and ANOVA analysis were used when appropriate to examine the statistical significance of the differences between groups of data. Statistical significance was placed at P < 0.05.
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