thrombin cleavage buffer - (Feb/13/2008 )

Hello guys,

I have a question regarding the thrombin cleavage buffer. Most of labs use Tris-NaCl (pH8) buffer with Ca as thrombin cleavage buffer. I am using this buffer to cleave my fusion protein too. But I notice that during cleavage reaction there is always some precipitant, which may bring down my wanted protein and lower the yield. I also notice that my protein is very soluble in PBS. Has anyone used PBS as thrombin cleavage buffer? Can I use PBS (pH 7.4) for the thrombin cleavage reaction? Is Calcium necessary for the thrombin activity? Thank you.

thrombin does not require any metals.