Cell surface Biotinylation
rudnick at biomed.med.yale.edu
Thu Mar 17 11:12:54 EST 1994
In article <ANITA.94Mar14111140 at piccolo.cco.caltech.edu> anita at piccolo.cco.caltech.edu (Anita Gould) writes:
>From: anita at piccolo.cco.caltech.edu (Anita Gould)
>Subject: Re: Cell surface Biotinylation
>Date: 14 Mar 1994 19:11:38 GMT
> From: rtfm at cix.compulink.co.uk ("Peter Smith")
> Organization: Abort, Retry, Fail Ltd
> Date: Sat, 12 Mar 1994 21:39:04 GMT
> Hi Folks,
> Does anybody out there have a good method for labelling cell
> surface molecules with biotin? I've been using 125I labelling with pretty
> good results, but my finger tips are getting fried with all that
> radiation! I'm sure that labelling the cell surface with biotin is
> possible....all I need is someone to tell me how!! Also any useful tips
> or pitfalls would be appreciated, I want to immunoprecipitate from the
> cells once they are labelled, will this be a problem?
> Thanks in advance from me and my finger tips!
>Yes -- some folks in my lab developed & published a biotinylation
>method. The ref is:
>Kayyem, JF et al, 1992. A method for the generation of monoclonal
>antibodies against rare cell-surface molecules. Eur J Biochem
>The only erratum/caveat I know of is that somewhere in the paper it
>mentions a step done at 4 deg, but it was actually done at 16 deg,
>because there's SDS in the buffer, & it ppts at 4 deg. (I'm pretty
>sure it was the avidin column step, which it sounds like you're not
>necessarily even going to be doing, unless you're going to go on to
>purify the cell-surface molecules).
>If you need any help or any further info, just drop me a line, & I can
>give you phone/fax/email addresses for people here to talk to.
> Best of luck!
> -Anita Gould
> anita at cco.caltech.edu
Here are two references that you could use for techniques:
1. Pietrini, G., Y. Suh, L. Edelmann, G. Rudnick and J. Caplan.
The axonal GABA transporter is sorted to the apical membrane of polarized
J. Biol Chem. 269:4668-4674 (1994).
2. LEVYTOLEDANO, R., L. CARO, N. HINDMAN and S. TAYLOR.
STREPTAVIDIN BLOTTING - A SENSITIVE TECHNIQUE TO STUDY CELL SURFACE PROTEINS
- APPLICATION TO INVESTIGATE AUTOPHOSPHORYLATION AND ENDOCYTOSIS OF
BIOTIN-LABELED INSULIN RECEPTORS.
ENDOCRINOLOGY. 133:1803-1808 (1993).
Gary Rudnick |voice (203) 785-4548
Department of Pharmacology |fax (203) 785-7670
Yale University School of Medicine |bitnet RUDNICK at YALEMED
333 Cedar Street, P.O. Box 208066 |internet:
New Haven, CT 06520-8066 |RUDNICK at BIOMED.MED.YALE.EDU
What separates the men from the boys is the price of their toys.
More information about the Methods