Biotin labelling of Cell Extracts for Western Blots

Martin Offterdinger a8803349 at unet.univie.ac.at
Tue Nov 12 10:25:51 EST 1996


On 8 Nov 1996 11:48:08 -0800, . at dub-mail-svc-1.compuserve.com,
@compuserve.com wrote:

>
>Hi All!
>I would like to be able to label cell lysates with biotin, immunoprecipitate
>the cell extracts with a specific antibody, do size separation on SDS-PAGE,
>and then detect bands on Western blots with horseradish peroxidase-conjugated
>avidin and ECL. I am hoping that this would circumvent using 35S-Methionine
>to label proteins in situ. Has anyone tried this ? I have not tried it yet, but once
>the pressure is off and I have some spare time (that's a laugh), I thought of trying
>it out. I would appreciate any suggestions to make the R&D time shorter.
>Thanks in advance.
>Martina
>TIMM at Hippo.Medcor.McGill.CA
>
Hi Martina!
 Labelling of proteins in situ with biotin works quite well but you
should consider the following: Biotinilation of proteins alters their
molecular weight as biotin itself has a MW of 0.6 kD and several
biotins are introduced into one protein molecule depending on which
labelling method you use. But you cannot simply neglect this MW
difference in many cases.



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