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Generation and characterization of anti-sulfoglucuronosyl paragloboside monoclonal antibody NGR50 and its immunoreactivity with peripheral nerve.

Abstract：

:Sulfoglucuronosyl paragloboside (SGPG) is a member of the sulfated glucuronic acid-containing glycolipid (SGGL) family found primarily in peripheral nerves. These glycolipids contain the HNK-1 carbohydrate epitope and are recognized by monoclonal IgM from patients with chronic demyelinating neuropathy and paraproteinemia. Recent studies indicate that SGGLs may serve as ligands for selectins, amphoterin, and laminin, suggesting that these glycolipids may play an important role in cellular adhesion. To elucidate the biological function of these glycolipids, we produced a murine monoclonal antibody (mAb) and studied its antigenic specificity. Using an enzyme-linked immunosorbent assay (ELISA), we found that the mAb designated as NGR50 belonged to the IgG2a subclass, and that the minimal titer (2 SD above the mean optical density value of control) of this mAb was 1:640, with 20 ng of purified SGPG as the antigen. Thin-layer chromatography (TLC) immunoblotting revealed that this mAb reacted specifically with SGPG and sulfoglucuronosyl lactosaminyl paragloboside (SGLPG), which is a structural analogue of the former, but not with other glycolipids. Desulfated derivates of SGPG and SGLPG did not react with mAb NGR50. Western blot analysis showed crossreactivity with human myelin-associated glycoprotein (MAG), but not with rat MAG or rat glycoprotein P0. Unlike anti-HNK-1 monoclonal antibody, however, NGR50 reacted only weakly with several proteins in the 20-30-kD regions, including human P0, suggesting that mAb50 has a different fine specificity as an anti-HNK-1 antibody. Immunocytochemical study of rat sciatic nerve using mAb NGR50 revealed positive staining at the outer surface of the myelin sheath and Schwann cells, as well as in the intervening connective tissues. Faint staining was also visible at the axolemmal-myelin interface; however, compact myelin was not stained.

journal_name

J Neurosci Res

journal_title

Journal of neuroscience research

authors

Yamawaki M,Ariga T,Bigbee JW,Ozawa H,Kawashima I,Tai T,Kanda T,Yu RK

doi

10.1002/(SICI)1097-4547(19960615)44:6<586::AID-JNR

subject

Has Abstract

pub_date

1996-06-15 00:00:00

pages

586-93

issue

eissn

0360-4012

issn

1097-4547

pii

10.1002/(SICI)1097-4547(19960615)44:6<586::AID-JNR

journal_volume

pub_type

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Generation and characterization of anti-sulfoglucuronosyl paragloboside monoclonal antibody NGR50 and its immunoreactivity with peripheral nerve.