Cell surface Biotinylation

Anita Gould anita at piccolo.cco.caltech.edu
Mon Mar 14 14:11:38 EST 1994


   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!

   Theresa

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
208:1-8. 

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



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