Help? Immunoprecipitation

Bernard Murray bernard at elsie.nci.nih.gov
Wed Feb 28 13:28:33 EST 1996


In article <313538C4.6FC5 at vision.postech.ac.kr>, pgs at vision.postech.ac.kr 
says...
>
>I am trying to discriminate some protein which have same 
>molecular weight with immunogloblin heavy chain.
>If anyone have tip that can solve this problem, please 
>tell me.
>Thanks in advance
>Myung Jong Kim
>My e-maol address is
>        pgs at vision.postech.ac.kr

The tricks I have used or heard used are;

1)	Treat your antibody to make it a Fab fragment.  This will reduce
	the size of the heavy chain.  The downside is that it can also
	reduce the affinity for antigen.
or
2)	Don't reduce your samples before electrophoresis (mercapto, DTT,
	DTE etc.).  The immunoglobulin will stay intact and run at a much
	higher weight.  The downside is that many proteins require
	reduction to give sharp bands.
or
3)	Covalently couple your antibody to a support (eg. CnBr Sepharose)
	then elute the antigen from the beads and run a normal gel.
	You get nice elution with eg. 2% formic acid.

There may be more ways.

		Good luck,
			Bernard

Bernard Murray, Ph.D.
bernard at elsie.nci.nih.gov  (National Cancer Institute, NIH, Bethesda MD, USA)




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