3 replies to this topic

[kedar]

Dear all,

here is the recipe for my loading buffer for western 6x.
tris.cl.
glycerol
sds
dtt
BPBlue
water

After keeping the 2x in 1.5 tubes for few weeks at 4 degrees, i can see the color has changed to dark green and precipitaes at the bottom. Is it still usuable? what's the problem?

thanks a lot for the replies in advance,

[mdfenko]

the sds has crystallized and the pH may have dropped slightly (absorbed co2).

warm the samples to room temperature (you can go higher) and mix to ensure that all of the sds has returned to solution.

if the sample remains green then you can either load it as-is and let the running buffer turn the bromphenol blue back to blue or you can add a little tris, pH 8, to the sample to raise the pH.

[kedar]

Hi, i heated and the precipitate disappeared but the color didn't change to blue. :-(

mdfenko, on 11 April 2011 - 07:47 AM, said:

the sds has crystallized and the pH may have dropped slightly (absorbed co2).

warm the samples to room temperature (you can go higher) and mix to ensure that all of the sds has returned to solution.

if the sample remains green then you can either load it as-is and let the running buffer turn the bromphenol blue back to blue or you can add a little tris, pH 8, to the sample to raise the pH.

[mdfenko]

then you can either load them as-is or add some tris to raise the pH slightly. there should not be a very noticeable effect on the gel either way.