Biomedical subjects

H Z Sheng

Publications and source records attributed to H Z Sheng.

23 records · Page 2Linked to original sources

PubMed1989

Developmental study of myelin basic protein variants in various regions of pig nervous system.

A developmental study of myelin basic protein (MBP) variants in eight regions of pig nervous system (NS) was performed using a quantitative electroimmunoblotting procedure. Four major MBP forms with apparent molecular weights of 21.5K, 20.2K, 18.5K, and 17.3K were identified in both the CNS and the PNS and were detected as early as 22 days before birth. Quantification of the most abundant forms, the 21.5K and 18.5K MBPs, revealed characteristic profiles of accumulation of these two variants in different regions of the NS. The ratio of 21.5K:18.5K MBP varied with developmental time as well as with the various NS regions, peaking 20 days postnatally. The 17.3K MBP was observed from embryonic stages to adulthood, as were the 21.5K and 18.5K forms. In contrast, the 20.2K variant appeared most abundant from 10 days before to 22 days after birth and thereafter decreased in intensity so as to be no longer detectable in the brain of a 5-year-old pig. A similar pattern was also observed with an anti-MBP-reacting protein with an apparent molecular weight of 23K. Taken together, these results suggest that in the pig NS, the expression of MBP variants may be regulated both regionally and developmentally.

Aging↗

PubMed1988

Electroimmunoblotting of small peptides separated on urea-dodecyl sulphate (SUDS) gels. Application to myelin basic protein.

A method for the electroimmunoblotting and immunodetection of peptides of less than 50 amino acid residues is described. Excellent resolution of a mixture of myelin basic protein (MBP) peptides was achieved by electrophoresis in a polyacrylamide stacking, urea-dodecyl sulphate minislab gel. Following electrophoresis, the peptides were transferred to various matrices and probed with monoclonal and polyclonal antibodies. Variables such as transfer time, membrane type, fixation and the amount of peptide loaded on the gel have been optimized as a consequence native and synthetic peptides can now be visualized in gels and immunodetected on immobilizing matrices. This procedure is particularly suited to the analysis and identification of small MBP fragments arising in various neuropathological conditions as well as for the rapid characterization of antigenic determinants recognized by monoclonal and polyclonal anti-MBP antibodies.

Animals↗

PubMed1988

Electroimmunoblotting of myelin basic protein peptides: a novel approach to the rapid characterisation of antigenic specificities of monoclonal and polyclonal anti-MBP antibodies.

A rapid and sensitive method for the identification of antigenic determinants recognised by monoclonal and polyclonal antibodies directed against myelin basic protein (MBP) is described. By electroimmunoblotting a series of overlapping peptides covering the entire MBP molecule with monoclonal anti-MBP antibodies, the binding pattern of immunoreactive peptides can be rapidly determined and the reactive antigenic determinant identified. This procedure, which can be performed with both native and synthetic peptides, can also with appropriate modification, be applied to the analysis of naturally occurring or experimentally induced polyclonal anti-MBP autoantibodies.

Antibodies↗

PubMed1987

Rapid characterization of protein epitopes recognized by monoclonal antibodies using direct probing on thin-layer and paper chromatograms.

A simple method for the comparison and identification of protein epitopes recognized by monoclonal antibodies directly on thin-layer plates and 3MM paper chromatograms is described. Enzyme digests of myelin basic protein were separated on thin-layer plates and 3MM paper, fixed with glutaraldehyde and probed directly with affinity-purified mouse monoclonal antibodies. Detection of the immunoreactive peptides was enhanced using a second rabbit anti-mouse immunoglobulin and finally located using an alkaline phosphatase-conjugated anti-rabbit immunoglobulin. By probing the same enzyme digests of MBP with various monoclonal antibodies raised against MBP, a different binding 'pattern' of reactive peptides is rapidly obtained for monoclonal antibodies of differing specificities. This procedure was extended to the identification of the antigenic determinant using synthetic peptides. The major advantages of this procedure are its simplicity, non-radioactive nature and speed. Furthermore, there is the possibility of sequencing immunoreactive peptides eluted from the 3MM paper.

Animals↗

PubMed1987

Use of protein-bearing nitrocellulose as immunogen for in vitro production of monoclonal antibodies: application to myelin basic protein electrophoretically separated from a complex brain protein mixture.

A procedure for producing monoclonal antibody to myelin basic protein (MBP) using in vitro immunization with MBP transferred to nitrocellulose is described. Following the separation of brain proteins by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and electrophoretic transfer of the electrophoretogram onto nitrocellulose, the MBP band located by immunodetection was excised from the nitrocellulose, ground, and used as immunogen for in vitro stimulation of unprimed mouse spleen cells. While in vitro immunization with soluble MBP was able to generate many hybrids, all the wells in the fusions carried out with the immobilized MBP contained hybrids, 33 to 42% of which were positive to MBP. Among these, six were further characterized; all were IgM and all bound to epitopes common to the 18.5K and 21.5K MBP forms of several species. In view of its simplicity, this technique should have a wide application for the rapid production of monoclonal antibodies to selected proteins or their fragments present in small quantity or difficult to purify on a large scale.

Animals↗