DEVELOPMENT OF A HIGHLY SENSITIVE CHEMICAL METHOD FOR DETECTING ALPHA-2-]8-LINKED OLIGO POLYSIALIC ACID RESIDUES IN GLYCOPROTEINS BLOTTED ON THE MEMBRANE/

A highly sensitive chemical method for detecting alpha 2-->8-linked ol igo/polysialic acid (oligo/polySia) chains was developed, including (i ) periodate oxidation, reduction with sodium borohydride, and subseque nt acid hydrolysis, giving rise to C-7 analogues and intact C-9 compou nds from nonreducing terminal and internal sialic acid residues, respe ctively; (ii) fluorescent labeling of these C-7 and C-9 compounds with 1,2-diamino-4,5-methylenedioxybenzene (DMB); and (iii) quantitation o f these DMB derivatives on fluorometric high-performance liquid chroma tography. As little as 1 ng of internal sialic acid residues of oligo/ polySia chains, the existence of which indicates the presence of oligo /polySia structure, was detectable by this method. This fluorometric C -7/C-9, analysis was successfully applied to glycoproteins blotted ont o a slit of polyvinylidene fluoride membranes and suggested the presen ce of some novel oligoSia-containing glycoproteins in pig embryonic br ains. (C) 1998 Academic Press.