fluorescein glycine amide: labelling
Dr Engelbert Buxbaum
engelbert_buxbaum at hotmail.com
Sun Aug 13 01:39:16 EST 2006
Raju Tatituri wrote:
> I am working with a hyperglycosylated lipid molecule and the MALDI mass is
> 25000-30000 and it looks like a big smear on the SDS-PAGE gel. But after
> labelling I can see the molecule as single protein band. I followed your
> protocol. I am unable to interpret my results.
Carbohydrate moieties in blycoproteins often contain acidic groups
(carboxylic acid, sulfonic acid), and since the sugar side chains are
not homogeneous the number of negative charges varies and hence
migration in SDS-PAGE. This results in the broad bands you observed.
Presumably some such carboxylic acids were modified by the reagent,
reducing charge inhomogenity and resulting in crisper bands.
Another way to get crisp bands with glycoproteins is to use the
positively charged detergent CTAB instead of the negatively charged SDS
(Anal. Biochem. 314 (2003) 70-76).
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