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protein extraction from fat breast tissue

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protein extraction from fat breast tissue

Postby Vero36 » Sat May 04, 2013 1:18 pm

Dear all,

I am a PhD student and I have a problem : we want to do quantitative proteomic on breast tumor and corresponding normal breast tissue. My problem is the following : normal bresat tissue is really fatty and the lipids interfere with my peptides quantification after trypsin digestion.

I use the micro BCA from Pierce. The problem is that it seems to overestimate the amount of peptides in normal breast tissue lysates compared to tumor lysates, and I think this is because of lipids that interfere with the microBCA dosage.

Would you have any idea to overcome this problem ?


Is there a trechnique to get rid of the lipids or is there a peptide quantification method that won't be wronged because of fat ?

Thanks a lot,

Vero
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Postby JackBean » Sat May 04, 2013 7:21 pm

I'd probably go for some sort of phenol-chloroform extraction which should effectively remove fats, but I'm not sure about how quantitative it is. But nowadays are available commercial kits.
What about to check publications to see what others do?
http://www.biolib.cz/en/main/

Cis or trans? That's what matters.
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