Concentration of dilute proteins for SDS-PAGE

HIRANYA ROYCHOWDHURY hroychow at NMSU.EDU
Mon May 6 10:00:03 EST 1996


On Fri, 3 May 1996, Joe Stains wrote:

> Does anyone know of any efficient way of concentrating large volumes of
> a dilute protein (several mls) for use with SDS-PAGE?  I have a couple
> of ideas on some techniques, but would be interested in anything anyone
> else may have, as they all have a lot of pros and cons!
> 
> Thanks in advance
> 
> Joe Stains
> 


Tha following are a few of the may ways, without the use of commercially 
avilable concentrators:

1. Precipitation with 90% ammonium sulfate- brings down almost 95% of the 
proteins. Dialysis against a low salt buffer required. 

2. Precipitation with acetone. The acetone conc. to be determined 
empirically.

3. Pptn with TCA. A part of the ppt. will not resolubilize, most proteins 
would dissolve back into Laemmli sample buffer and the resulting SDS-PAGE 
pattern would be fairly representative.

4. EtOH pptn. works the same way as the TCA pptn., and is less harsh. 
But, a good percentage of the proteins may not ppt. in alcohol.

5. PEG 8000. This is my favorite. But, the salt concentration also 
increases so one may need to carry out drop dialysis before preparing 
sample for electrophoresis.



			>>>>>>>>>>>>>>>>>>>>>>>>>>>
			  Hiranya S. Roychowdhury
   			  Plant Genetic Engineering Lab.
			  Box 3GL, NM State Univ.
			  Las Cruces, NM 88003
			  Phone: (505) 646-5785
			  hroychow at nmsu.edu
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