I have problems in my last western blots performed with samples of mice liver: I observed only a strong signal underlining the band of SR-BI protein (my target), but the control protein is not detected (weak β actin signal and/or spot not defined as well as not uniform in all the samples). First I used the anti-β actin antibody, and in other experiments I incubated the liver samples with other antibodies for other proteins (such as GAPDH) in order to find a reliable control protein to compare the SRBI signals; despite these changes, I obtained the same results.
My question is: knowing that the samples have been defrosted and re-used numerous times, is it possible that the samples are starting to be degraded, in particular the low-weight proteins (β actin, GAPDH) instead higher weigh proteins, such as SRBI (75kDA ca.)?
Help me pleaseee !