The species distribution of nervous system antigens that react with anti-myelin-associated glycoprotein antibodies

The reactivity of monoclonal and polyclonal antibodies directed against human central nervous system (CNS) myelin-associated glycoprotein (MAG) was investigated in a number of animal species. The antibodies included mouse monoclonal antibodies obtained by immunization with human MAG; HNK-1, a mouse monoclonal antibody raised against a human lymphoblastoma and used to identify a subset of lymphocytes with natural killer function; human IgM paraproteins associated with neuropathy; and polyclonal antibodies obtained from rabbits immunized with rat or human MAG. Following polyacrylamide gel electrophoresis of CNS and peripheral nervous system (PNS) tissue from human, bovine, cat, rabbit, guinea pig, rat, mouse, frog, gold fish and chicken, proteins were electrophoretically transferred onto nitrocellulose. The immune-staining of electroblots showed distinct interspecies variation in the reactivity of the antibodies with MAG. In addition, the species distribution of several low molecular weight glycoproteins present in PNS tissue that cross-react with anti-MAG antibodies was determined. These low molecular weight antigens are not present in CNS homogenates or in purified human CNS myelin. It was also shown that IgM from a patient with peripheral neuropathy and paraproteinemia associated with anti-MAG antibodies recognized these low molecular weight antigens. The results suggest that IgM paraproteins, HNK-1 and some mouse monoclonal antibodies react with carbohydrate determinants shared by MAG and several lower molecular weight glycoproteins present only in human, bovine, cat and chicken PNS. Rabbit polyclonal anti-rat MAG antisera and mouse monoclonal antibodies reacting with peptide epitopes of MAG are much more specific for detecting MAG than antibodies reacting with carbohydrate epitopes of human MAG. The results are discussed in relation to human demyelinating peripheral neuropathy associated with IgM paraproteinemia

Myelin-associated glycoprotein and related glycoconjugates in developing cat peripheral nerve: a correlative biochemical and morphometric study

The expression and accumulation of the myelin-associated glycoprotein (MAG) and other glycoconjugates have been studied during myelination in the developing cat peripheral nervous system. The glycoconjugates studied have in common a similar carbohydrate determinant which is bound by many antibodies, including the mouse monoclonal antibody HNK-1, and human IgM paraproteins from patients with neuropathy. In addition to MAG, the reactive glycoconjugates include a 60-kilodalton (kD) glycoprotein and a group of 20-26 kD glycoproteins, as well as a group of recently identified acidic glycolipids, the major one of which is sulfate-3-glucuronyl paragloboside (SGPG). The accumulation of these glycoproteins and glycolipids is compared with the established myelin proteins P0, P1, and P2 and with morphometric indices of myelin volume and axonal perimeter. The study demonstrates that MAG appears and accumulates very early during myelination, being present at 15% of the maximum level prior to the appearance of P0, and at 80% of the maximum level when P0 is at 30% of its maximum level. In the adult, the level of MAG falls to 60% maximum. The 60 kD and 20-26 kD glycoproteins accumulate at the same time as or later than P0, suggesting that they are either compact myelin proteins or in membranes closely associated with compact myelin. SGPG accumulates with P0 early in myelination, but falls to 60% of maximum in the adult. By comparing biochemical and morphometric data, we demonstrate that P0 and other compact myelin proteins accumulate synchronously with the increase in myelin area. MAG accumulation, however, is closely related to changes in axonal perimeter, consistent with a predominant localization of MAG to the periaxonal membranes in the peripheral nervous system

Local Mobility in Lipid Domains of Supported Bilayers Characterized by Atomic Force Microscopy and Fluorescence Correlation Spectroscopy

Fluorescence correlation spectroscopy (FCS) is used to examine mobility of labeled probes at specific sites in supported bilayers consisting of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) lipid domains in 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC). Those sites are mapped beforehand with simultaneous atomic force microscopy and submicron confocal fluorescence imaging, allowing characterization of probe partitioning between gel DPPC and disordered liquid DOPC domains with corresponding topography of domain structure. We thus examine the relative partitioning and mobility in gel and disordered liquid phases for headgroup- and tailgroup-labeled GM1 ganglioside probes and for headgroup- and tailgroup-labeled phospholipid probes. For the GM1 probes, large differences in mobility between fluid and gel domains are observed; whereas unexpected mobility is observed in submicron gel domains for the phospholipid probes. We attribute the latter to domain heterogeneities that could be induced by the probe. Furthermore, fits to the FCS data for the phospholipid probes in the DOPC fluid phase require two components (fast and slow). Although proximity to the glass substrate may be a factor, local distortion of the probe by the fluorophore could also be important. Overall, we observe nonideal aspects of phospholipid probe mobility and partitioning that may not be restricted to supported bilayers