A protein question******

lamison at esvx12.es.dupont.com lamison at esvx12.es.dupont.com
Mon Apr 25 10:11:43 EST 1994


>We have 11 ug of a protein isolated from polyacrylamide gel
>electrophoresis. We are interested in isolating the protein from the gel
>and making antibodies to it....

I would just inject the gel into the animal.  After the gel is stained and
destained, cut out the band of interest and wash for ~8 h with many changes of
PBS/TBS to remove the destain components and SDS.  Then lyophilize (~24h) and
grind to a very fine powder.  Rehydrate with a minimal amount of PBS/TBS
(enough for a slurry and at standard immunization protocol protein 
concentration).  Immunize as usual except no need for adjuvant as the gel
itself does an adequate job. Make sure the gel powder is ground fine enough to
pass through the syringe needle.

I have used this method many times for antibody to denatured SDS-PAGE purified
protein for use in Western blots with good success and strong antibody.



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