The protein isoforms I am interested in comparing appear as distinct bands on the gel I have already run. I have an Excel sheet with optical density measurements I obtained using ImageJ; it looks something like this:
Lane Iso1 Iso2 GAPDH 1 149.06 194.646 893.08 2 832.654 494.473 148.335 3 49.998 490.539 147.361 4 29.347 208.53 120.652
For the other proteins I've analyzed so far, I've been computing fold-change relative to my loading control, GAPDH, with the following equation:
Fold-Change = Log2(Protein/GAPDH)
I'm not sure how best to compare my isoforms to each other and considering the following two equations:
1. Fold-Change = Log2[(Iso2/GAPDH)/(Iso1/GAPDH)] 2. Fold-Change = Log2(Iso2/Iso1)/GAPDH
I've already computed both of these values for all my samples, graphed them and found that the resulting graphs look pretty different. Which of the two equations do you think is a better way to compare the relative quantities of these proteins?
Alternative equations are also welcome.