Semi-quantitative westerns

[MedGen]

I'm currently doing some immunoblots and quantifying them using integrated densitometry.

 

I've probed for my protein of interest and B-actin as the control housekeeping protein. I've also normalized the bands against the background signal.

 

I want to combine the data from three separate membranes so that I can statistically analyse them (I'm being a little naughty, but I need to be able to confirm a previous finding from this). So I've run the blots in triplicate, and normalised the bands to the background.

 

The question is whether I can use the relative ratio of b-actin to protein of interest as a general (i.e. semi-quantitative measure) of expression or whether I have to retain the integrated density as the measure of expression?

 

Thanks in advance.

 

p.s. the integrated density measurements are done in Adobde Photoshop CS4 using 1200 dpi 16-bit grey scale TIF files scans of the developed membranes.