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adjusting pH of 4x stacking buffer


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#1 pinkc

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Posted 14 August 2012 - 11:36 AM

I am following a protocol that requires the stacking buffer be adjusted to a pH of 6.8. I accidentally overadjusted it to around 6.1 so I used some NaOH to compensate the pH back to 6.8. Would this affect the stacking gel performance or not really?

#2 bob1

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Posted 14 August 2012 - 01:13 PM

The NaOH adds Na to your solution (the OH- combines with the H+ in the solution to form water), if the assay is dependent on salt you may have a problem, but it should be fine most of the time.

The usual way to do this sort of thing is to make a tris solution of pH 6.8 and add that to your stacking buffer.

#3 mdfenko

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Posted 15 August 2012 - 05:07 AM

the addition of naoh will affect the stacking buffer.

if you overshoot then you should add more basic component (tris), calculated for a greater volume, readjust pH, then bring to the new final volume.
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#4 pinkc

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Posted 16 August 2012 - 11:16 AM

Ack! OK thanks a lot for replying - I haven't used it yet but now I will dump it out and remake it and be careful about adjusting pH.




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