Then, it was incubated with anti-rabbit or anti-goat secondary antibodies conjugated with horseradish peroxidase (1:1000) and finally detected by chemiluminescence and autoradiography with x-ray film. Negative controls were obtained in the absence of the primary antibody. The density of each band was normalized to the density of the actin B band that was used as an internal control.
Quantification for Western Blot Assay
In each experiment, equal amounts of protein were loaded for all samples, and both groups in one experiment were loaded on the same gel. For quantification, a screening was performed on blots with x-ray film using different times of exposure to optimize the signal. The levels of protein were compared and analyzed with densitometric studies by Scion Image for Windows. Optical density data are expressed as arbitrary units 6 SEM (n = 8).
Data are expressed as the mean 6 SEM. Representative gels are shown in the figures. Statistical analysis was performed by the paired or unpaired ?-test. Values of P < 0.05 were considered significant.