Labeling CLIP-tag Purified Protein In Vitro (P9311)

Protocol

1. Thaw the CLIP-tag Purified Protein or CLIP-tag fusion protein at room temperature just before use.
   
   
2. Set up the reactions, in order, as follows:

   Component	Volume	Final Concentration
   Deionized Water	32 μl
   5X CLIP-tag Reaction Buffer	10 μl	1X
   50 mM DTT	1 μl	1 mM
   50 µM CLIP-tag Purified Protein	5 μl	5 µM
   250 µM CLIP-tag Substrate	2 μl	10 µM
   Total Volume	50 μl

3. Incubate in the dark for 60 minutes at 37°C. 
   
   
4. Run sample on an SDS-PAGE gel and detect using a fluorescent gel scanner or store samples at -20°C or -80°C in the dark.
   
   
   Notes for In Vitro Labeling
   Adjust the volumes to achieve the final concentrations indicated if the stock solutions of protein, substrate, and DTT are different than described above. 
   
   One may choose to label the CLIP-tag Purified Protein or CLIP-tag fusion protein in a different final concentration, however, it is recommended to use a 2-fold excess of CLIP-tag substrate vs. CLIP-tag protein. 
   
   Labeling may also be carried out in 1X PBS solution or 50 mM HEPES containing 1 mM DTT.