[ccp4bb] Offtopic - In vitro protein interaction analysis using Diazirine crosslinkers.

[Rohan Agashe]

Apologies for an off topic question. I was hoping for some advice regarding the use of sulfo-NHS diazirine crosslinker (Pierce - http://www.piercenet.com/products/Browse.cfm?fldID=DB013E61-5056-8A76-4EC4-473A6F2468A1) for in vitro protein interaction analysis. Have tried labeling the protein (9kDa, 10 to 20uM conc) with 2 primary amines using 30-40X excess Sulfo-NHS-Diazirine crosslinker in PBS (1hour at 4deg). Have tried other concentration combination as well, but this ratio yields some usable material. However, majority of the protein crashes out of solution. Was wondering if anyone has any suggestions in optimizing the yield of crosslinker labelled protein? Thanks in advance. regards, Rohan Agashe