Adding Neutravidin to CM5 chip

I would like to couple a CM5 chip with Neutravidin, block with Ethanolamine and then add my biotinylated protein (looking at protein-peptide binding). Has anyone had experience with this setup? I guess I am interested in knowing how well it worked for others and what amount of Neutravidin they have had good luck with.

Any other hints/suggestions would be greatly appreciated.

Thanks!
Claude Bessy

Hi Claude

I don't have a perfect answer for you but some references to start with.

I would start with a normal amine immobilization and culculate what desnity you want.
As an alternative you can use the SA-sensorchip with straptavidin on the surface. That will speed up your experiment.
You can also use the CAP-regaent sensor chip. This is a more flexible way of immobilization using the biotin-avidin system. Take a look!


O'Shannessy, D. J., Brigham-Burke, M., and Peck, K.; Immobilization chemistries suitable for use in the BIAcore surface plasmon resonance detector. Anal.Biochem. (205): 132-136; 1992.
Gitlin, G., Bayer, E. A., and Wilchek, M.; Studies on the biotin-binding sites of avidin and streptavidin. Tyrosine residues are involved in the binding site. Biochem.J. (269): 527-530; 1990.

Wilchek, M. and Bayer, E. A.; Introduction to avidin-biotin technology. Methods Enzymol. (184:5-13.): 5-13; 1990.

best regards

Arnoud

Thanks Arnoud!

Sounds like this is going to be very experimental. I was hoping to reduce costs by manually adding the avidin (I know, this will probably haunt me in the end).

I will post my findings for someone to use in the future.