protein pptn in acetone problem

Ron Tate rtate at bmb-fs1.biochem.okstate.edu
Mon Nov 20 17:27:47 EST 1995


brett at BORCIM.WUSTL.EDU (brett) wrote:
>Hello, I'm trying to ppt. my protein using cold acetone. It was in a 50uM
>sodium phosphate buffer. I added 8 vol. acetone, and now have a large,
>chunky ppt. I believe the large chunks are phosphate, as buffer alone gave
>the
>same appearance. Parallel ppt's of the same protein in a citrate buffer
>gave a finer, milky ppt. I now want to dry down the pellets and resuspend
>them for SDS-PAGE analysis, but am worried about the salt. Anyone have any
>suggestions for what to do? I thought I might resuspend the chunky pellets,
>dialyze into another buffer, and reppt. Thanks in advance,
>
>
>
>Brett Lindenbach
>    
>Program in Immunology                              
>Washington University - St Louis                  
>brett at borcim.wustl.edu                             
>
>"I own my own pet virus. I get to pet and name her." - Cobain
>

 I have successfully precipitated protein out of a 20mM potassium phosphate buffer for 
SDS-PAGE.  I add TCA to 10% final concentration and let it sit for 10-20 minutes collect by 
cetrifugation and wash the pellet with acetone.  This has given some nice gels from pretty 
dilute samples.

Happy Hunting
Ron Tate
OSU BMB




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