ON THE SPECIFICITY OF ANTI-SULFOGLUCURONOSYL GLYCOLIPID ANTIBODIES

The mouse monoclonal anti-human HNK-1 antibody (also variously known a s L2, Leu-7, CD57, VC1.1), monoclonal anti-sulfoglucuronosyl glycosphi ngolipids (SGGLs) antibody (mAb NGR50), and human sera from patients w ith demyelinating neuropathy and IgM paraproteinemia, are known to rea ct with not only SGGLs, including sulfoglucuronosyl paragloboside (SGP G) and sulfoglucuronosyl lactosaminyl paragloboside (SGLPG), but also glycoproteins, such as myelin-associated glycoprotein (MAG), P0, PMP22 , and certain adhesion molecules. These antigens are known to possess the so-called HNK-1 epitope (3-sulfoglucuronic acid, SGA) moiety. To f urther define the precise structural requirement of this carbohydrate epitope, we chemically synthesized 14 SGGLs, and their nonsulfated der ivatives with defined carbohydrate chain lengths and aglycone structur es. The various aglycones include ceramide, 2-(tetradecyl)hexadecyl (B 30), and 2-(trimethylsilyl)ethyl (SE). These synthetic SGGLs were test ed for their immunoreactivity with the above antibodies by high-perfor mance thin-layer chromatography (HPTLC)-immunoblotting and ELISA. The anti-HNK-1 antibody (VC1.1) reacted with SGGL analogues containing a m inimum of two sugars (SGA-Gal-Cer), but not with non-sulfated derivati ves of SGGLs nor with SGGLs having a modified ceramide structure. mAb NGR50, on the other hand, reacted with only SGPG and SGLPG. A human pa tient serum (LT) reacted with all synthetic SGGLs except those with an SE aglycone structure. On the other hand, another human patient serum (YT), like the anti-HNK-1 antibody, VC1.1, reacted with SGPG, SGLPG, and SGGL analogues containing a minimum of two sugars (SGA-Gal-Cer). A ll antibodies reacted more strongly with synthetic SGGLs with longer c arbohydrate chains. These results indicate that anti-SGGL antibodies r ecognize a minimum of two sugars bearing the following structure (3-su lfoglucuronosyl beta 1-3 galactosyl, SGA-Gal-) and that the aglycone ( ''ceramide'') structure appears to play an important role for antibody -antigen interaction.