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-   -   Western Blot: Actin bands too thick? (http://www.molecularstation.com/forum/western-blot-forum/87692-western-blot-actin-bands-too-thick.html)

ajc613 03-05-2013 06:17 PM

Western Blot: Actin bands too thick?
 
New to western blotting, and trying to figure out the best way to do the loading control using beta actin. Because my protein of interest is not expressed at high levels, I am loading a lot of total protein in each lane to get a good signal. But that makes the actin bands really large. Is there a way around that?
Thanks!

scottishov 06-16-2013 04:16 PM

Re: Western Blot: Actin bands too thick?
 
Quote:

Originally Posted by ajc613 (Post 446809)
New to western blotting, and trying to figure out the best way to do the loading control using beta actin. Because my protein of interest is not expressed at high levels, I am loading a lot of total protein in each lane to get a good signal. But that makes the actin bands really large. Is there a way around that?
Thanks!

Are your doing colorimetric or chemiluminescence? If its the latter then just expose for a shorter period to get smaller bands and hence see any differences. Either that or use your antibody at a lower concentration. Probably the secondary over the primary in my experience.


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