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MS-compatible methods for blocking agarose beads


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#1 treesandcars

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Posted 10 October 2013 - 04:39 PM

I'm trying to IP my protein of interest and identify interactors by mass spec. By silver staining I see tons of nonspecific binding in both my IP and my negative control IP.

 

Increasing the stringency of my washes doesn't help, but pre-blocking my agarose beads with BSA gets rid of most of these nonspecific bands. Unfortunately, this isn't compatible with mass spec because 1) my protein of interest runs similarly to BSA and 2) I don't want to fill the ion trap with BSA peptides.  

 

Any suggestions on ways to block agarose beads that are compatible with mass spec?



#2 neuron

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Posted 18 October 2013 - 02:03 AM

BSA is just a protein, which is used for blocking everywhere in western blotting etc. Can't you use any other small molecular weight protein for blocking?  Since BSA is standard, you can use some other standard protein like lysozyme etc. 






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