Whole-cell lysate samples were immediately frozen after treatment completion. Membrane/cytosol fractionated lysates were processed immediately after the treatment without a freezing step. Further details of homogenization and western blotting technique can be consulted in Cilleros-Mañé et al. [41 (link)].
The densitometry of the bands was obtained with ImageJ software. The integrated optical density of the bands was normalized with respect to: (1) the background values; and to (2) the total protein transferred on PVDF membranes, measured by total protein analysis (Sypro Ruby protein blot stain, Bio-Rad [49 (link)]). The relative variations between the experimental samples and the control samples were calculated from the same membrane image. All presented data derive from densitometry measurements made of three to ten separate replicates, plotted against controls. Data quantification was performed blindly.
The densitometry of the bands was obtained with ImageJ software. The integrated optical density of the bands was normalized with respect to: (1) the background values; and to (2) the total protein transferred on PVDF membranes, measured by total protein analysis (Sypro Ruby protein blot stain, Bio-Rad [49 (link)]). The relative variations between the experimental samples and the control samples were calculated from the same membrane image. All presented data derive from densitometry measurements made of three to ten separate replicates, plotted against controls. Data quantification was performed blindly.
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