On the specificity of anti-sulfoglucuronosyl glycolipid antibodies

The mouse monoclonal anti-human HNK-1 antibody (also variously known as L2, Leu-7, CD57, VC1.1), monoclonal anti-sulfoglucuronosyl glycosphingolipids (SGGLs) antibody (mAb NGR50), and human sera from patients with demyelinating neuropathy and IgM paraproteinemia, are known to react with not only SGGLs, including sulfoglucuronosyl paragloboside (SGPG) and sulfoglucuronosyl lactosaminyl paragloboside (SGLPG), but also glycoproteins, such as myelin-associated glycoprotein (MAG), PO, PMP22, and certain adhesion molecules. These antigens are known to possess the so-called HNK-1 epitope (3-sulfoglucuronic acid, SGA) moiety. To further define the precise structural requirement of this carbohydrate epitope, we chemically synthesized 14 SGGLs, and their nonsulfated derivatives with defined carbohydrate chain lengths and aglycone structures. The various aglycones include ceramide, 2-(tetradecyl)hexadecyl (B30), and 2-(trimethylsilyl)ethyl (SE). These synthetic SGGLs were tested for their irnmunoreactivity with the above antibodies by high-performance thin-layer chromatography (HPTLC)-immunoblotting and ELISA. The anti-HNK-1 antibody (VC1.1) reacted with SGGL analogues containing a minimum of two sugars (SGA-Gal-Cer), but not with non-sulfated derivatives of SGGLs nor with SGGLs having a modified ceramide structure. mAb NGR50, on the other hand, reacted with only SGPG and SGLPG. A human patient serum (LT) reacted with all synthetic SGGLs except those with an SE aglycone structure. On the other hand, another human patient serum (YT).