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Where does mouse BRCA1 migrate on a western blot?

BRCA1 MOUSE Western Blot

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#1 Epigeneticist

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Posted 28 September 2013 - 03:44 PM

Does anyone have experience and success with detecting mouse BRCA1 by western blot? BRCA1 antibody datasheets indicate the protein should be around 220kd. I feel this specific to human because I think mouse is actually a smaller protein. I feel that mouse BRCA1 migrates around 200-210kd because I a using a new antibody with less background and the predominant band is around 205kd. Most antibodies produce a lot of non-specific bands so I am trying to figure out which band is correct. I have knocked down BRCA1 but with so many non-specific bands and not knowing where mouse BRCA1 migrates, it is hard to determine which band is correct. I also load at least 50ug protein fro whole cell lysate and have used nuclear lysates too. 

 

Thanks!



#2 bob1

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Posted 30 September 2013 - 11:47 PM

The classic way to do this would be to get some peptide against the epitope of the antibody and then pre-incubate the antibody with the peptide.  Finally use the pre-incubated antibody to probe a membrane, the abolished bands should be the specific ones.

 

However, I know that BRCA1 is particularly problematic with respect to background in humans, and I haven't tried it in mice, so can't really help there.







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