Tricks to eliminate non-specific binding of proteins to amylose resin, any ?

Artem Evdokimov AEVDOKIMOZ at cinci.rr.com
Thu Jan 17 22:57:38 EST 2002


Hi,

You do not specify what protein(s) you work with. There is a multitude of
reasons as to why the protein fusions may 'stick' - you'd be well advised to
provide more details before we could propose anythig but pure speculation :)

Normally, MBP fusions are run in 25 mM phosphate pH 8.0, 300 mM NaCl which
is the recommended buffer as far as manufacturer is concerned. Adding
ethylene glycol or glycerol often helps and so does the addition of nonionic
detergents.

A.G.E.

""Khadka, Deepak (NICHD)"" <khadkad at mail.nih.gov> wrote in message
news:000C316550E62144830CDFCEDA73050701E6282E at nihexchange4.nih.gov...
>
> I am performing binding assay of some test proteins with MBP-fusion
protein.
> When I try to purify the mixture by using amylose resin, most of the test
> proteins gets stuck to the resin. My buffer (both for binding and washing)
> is 50 mM PIPES (pH 7.0), 100 mM NaCl and 1 mM EDTA. Playing with the salt
> concentration and adding BSA did not help. Any suggestions ?
>
> Deepak
> NICHD/NIH
>
> ---





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