Yellow Laemmli Sample Buffer - (Apr/02/2008 )
I have some laemmli sample buffer which is yellow, the pH is too low.
What are the problems with the buffer being too low?
migration troubles.
Usually the migration buffer is well buffering, and when you load your sample it turns blue again. Then it's fine.
Depends on what you did to get a yellow sample buffer.
The pH is too low, I never adjusted TrisHCl solution, just added the correct moles.
I thought I read, it may cause protein degradation or something. Makes sense though that when you load it the running buffer would buffer it back to a good pH. If that's the case then I'll use it 
What are the problems with the buffer being too low?
do you work with TCA-precipitated samples? or the sample buffer was spoiled (with acid), or was wrongly mixed
No, no, the sample buffer's not spoiled. Instead of using a stock solution of TrisHCl that is adjusted to a certain pH, I just added the solid TrisHCl (with all the other ingredients). I never adjusted pH. I know I could adjust pH at this point.
But my question is about the effects of the yellow (low pH) Laemmli?
you should correct the pH to ensure that your samples run correctly (in fact, i would throw the buffer out and make it fresh).
I want to know why you would throw it out. What is bad about such a buffer?
So, to clarify, the question is - what would the low pH do to the protein samples?
I want to know why you would throw it out. What is bad about such a buffer?
So, to clarify, the question is - what would the low pH do to the protein samples?
the local excess of protons of your sample may be buffered by the gel specific buffer or running buffer; I think you may try particularly if a new preparation takes pain...
So what would low pH do to protein samples.
acidic pH may denaturize proteins; harsh pH is used to precipitate proteins, f.i. by TCA or formic acid; SDS-PAGE sample buffer should solve acid precipitated proteins; check if your proteins are solved after heating in sample buffer