Protein formed ppt after stored at -80oC - (Sep/01/2006 )

Dear All,

I purified my overexpressed protein from E.coli by His column yesterday. I kept the protein at
-80oC fridge. Today, I got it back from the fridge. After I thawed the protein. There were some ppt and these ppt cannot be dissolved.

Do anyone have this experience? How to dissolve them?

thx very much!!!!

siuchi

well, i've nerver faced that, but noticed that BSA forms pellets that are removed by 25° heating.
You may heat an aliquot of your sample (containing some of the ppts) gently and see what happens

Were your proteins only in PBS? If they are freezed in PBS without glycerol and they are too much concentrated, they might precipitate.

the protein may have been denatured by ice crystal formation. you may be able to suspend it in denaturing conditions (eg- sds, urea, etc) but it would not regain function.