Quantification of Sugars on glycoproteins

John Proudman, GGPL, B-262 johnp at GGPL.ARSUSDA.GOV
Thu May 11 09:24:28 EST 1995


Oops, sorry Frank, I realized after I sent this that I had not focused on the 
"Western blot" part of your question!  I shouldn't read my EMail before my 
second cup of coffee.  Obviously, this will not work for a Western.  I think 
quantification with either the Oxford or Boehringer kit for Westerns would be 
quite relative unless you have a purified standard of the same protein.

John
>
>Frank:
>You might try the "DIG Glycan ELISA" sold by Boehringer Mannheim (Cat.
>#1460366).  You adsorb your glycoprotein to the wells of an ELISA plate,
>incorporate DIG into the CHO by oxidation, and quantify the DIG relative
>to a fetuin standard.  The standard curve goes down to 2 ng of fetuin.
>In my limited experience, the standard curve works very well and the
>sensitivity is as advertised.  How well it detects your oligosaccharide
>may depend on the amount of CHO present and perhaps its tertiary
>structure (accessability of the CHO to reagents).
>
>John
>
>John A. Proudman
>Germplasm & Gamete Physiology Laboratory
>Beltsville Agricultural Research Center
>Agricultural Research Service
>U.S. Department of Agriculture
>Beltsville, MD 20705 USA



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