Protocol Online logo
Top : New Forum Archives (2009-): : SDS-PAGE and Western Blotting

Cell Lysis Buffer - (Oct/29/2010 )

Can someone give me a recipe for a cell lysis buffer...here is what I"m trying to do:

Detect Ras in HEK293 cells by western.

Thanks!

-bioJ-

1% Triton x-100
50 mM Tris pH7.4
150 mM NaCl
1 mM EDTA
Protease/phosphatase inhibitors

Should do the trick.

-madrius1-

great thanks! What is the difference between using tritonx or sds as the detergent?

-bioJ-

I think it is their degree of "harshness", whatever that means. What method are you using for protein quantitation?

-Halfro22-

triton is non-ionic and sds is ionic

-mdfenko-

1X Cell Lysis Buffer:
20 mM Tris-HCl (pH 7.5)
150 mM NaCl
1 mM Na2EDTA
1 mM EGTA
1% Triton
2.5 mM sodium pyrophosphate
1 mM b-glycerophosphate
1 mM Na3VO4
1 μg/ml leupeptin
some protease inhibitor added will make it better~

-castleinthesky-

castleinthesky on Fri Mar 2 03:07:09 2012 said:


1X Cell Lysis Buffer:
20 mM Tris-HCl (pH 7.5)
150 mM NaCl
1 mM Na2EDTA
1 mM EGTA
1% Triton
2.5 mM sodium pyrophosphate
1 mM b-glycerophosphate
1 mM Na3VO4
1 μg/ml leupeptin
some protease inhibitor added will make it better~

Castleinthesky, it might be worth you checking when these posts were last posted in, this one is well over a year old...

-bob1-