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Western Blot Forum Discuss western blotting and immunoblot in the western blot forum discussion board. Ask Questions about transfers, blocking, membrane antibody incubation and exposures.


Western blotting of brain proteins

Western blotting of brain proteins - Western Blot Forum

Western blotting of brain proteins - Discuss western blotting and immunoblot in the western blot forum discussion board. Ask Questions about transfers, blocking, membrane antibody incubation and exposures.


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Old 04-06-2011, 05:18 PM
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Default Western blotting of brain proteins



I'm a cancer biologist who has been running westerns for 10 years without any major problems. I have recently started trying to run westerns of brain proteins. Brains were snap-frozen, sectioned, proteins extracted using RIPA and run on either a 4-12% Bis-tris gel or a 12% bis-tris gel. The bands we are getting are totally weird. Within single lanes, they look like a caret shape (i.e. rather than having nice flat horizontal bands, they look like an upside down V ). Sometimes it is really bad where the band is almost completely vertical. We have tried altering gel concentration, running the gels slower for longer (i.e. around 100V for 1.5 hours), putting the gel boxes in an ice bucket, etc.

Does anyone have an solution to this problem. I have heard that brain protein is very salty due to a high ion content and this can interfere with westerns but I am not sure what to do about this. Any suggestions would be appreciated
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