UPDATE: the temperature sensor of the pH meter was broken

Hello,

I use this buffer during my protein purification process:

20mM Tris-HCl pH 8.0, 6M urea, (+ protease inhibitors)

In one step of the protocol, I have to lower the pH to 2.5 (my protein is soluble at this pH), ultracentrifuge, and then raise the pH to 8.5 again. I use HCl and NaOH to adjust the pH and I do it on ice. After a while, the pH drops from 8.5 to around 2.5-4.

I did a test and took buffer without a protein and measured it at two temperatures. On ice, it was ~3.5 and at 25C it was 8.5. Is there something wrong with the buffer or the pH electrode?

I just can add that my colleague had a similar problem with Tris buffer at pH 7.5.

Thank you :)