Hi
I mixed Tris HCl, EDTA, NaCl and deoxycholate 0.5% to make up a 1x RIPA buffer stock. After overnight in fridge, I found that my buffer becomes gel-like! It literally form a 'jelly' in my reagent bottle.
Does anyone have similar experience and can share the possible cause of this?
It occurs due to protein denaturation and aggregation (clumping) from heat. To fix this, make sure to pre-chill equipment. Sonication produces intrinsic heat, so be weary. You may also want to increase the volume of lysis buffer, however, this is at the consequence of a lower protein concentration.
The solution I recommend is to pass your samples through a 27-gauge needle until the solution becomes less viscous. Bubbles or foam may form, but centrifuging helps.
Hi Zunair and Stjepan
The recipe I used was 10 mM Tris pH 7.5, 100 mM NaCl, 2 mM EDTA and 0.5% deoxycholate. There was not other detergent.
The buffer forms a gel-like itself so I wasn't even able to pipet it into my samples. On vigouros shaking the gel 'dissolved' to become viscous liquid. Is this normal for deoxycholate-containing buffers like RIPA?
The culprit is deoxycholate
Article Deoxycholate-hydrogels: Novel drug carrier systems for topical use
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