I have frozen a double-purified recombinant protein (118 kDa) in HEPES buffer before the addition of 50% glycerol, by mistake.
How damaging can this mistake be for the activity of the protein according to your experience?
Thanks Paul,
after having realized the mistake, I immediately thawed the protein and added the glycerol 50%. I just hope it's still functional, I'll test it during the upcoming days!
Freezing time also matters, If we freez slow, then water particles get time to make big crystals and basically these crystals harm the protein; or cells. But Believe me! only one time freezing will not harm proteins (different for cells, as cells are more sensitive.). I will suggest you to make aliquotes; to lessen the freez-thaw cycles. It really matters.
In my experience, some proteins don't care at all if glycerol is present. Others totally need glycerol for stability. And it also depends sometimes on the concentration of protein. So if you have an activity assay, I would not assume anything. I would empirically try to determine the best conditions for freeze/ thawing.
- Badges
- Science method