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The 21,500 molecular weight (21.5K) variant of myelin basic protein (MBP) was isolated from sheep brain and partially characterized. Digestion with cyanogen bromide and trypsin yielded peptides which showed that approximately 30 additional amino acids were inserted at the equivalent of the amino acid at position 57 in the bovine 18.5K MBP sequence. An unusually hydrophobic peptide Pro, Val, Leu, Trp, Lys was present in this region. Ornithine was present in hydrolyzates of 21.5K MBP, but it was not detected in any of the peptides.

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.

A solid phase competitive assay for human myelin basic protein (MBP) has been developed using a monoclonal antibody to MBP. The assay has been applied to the detection of antigenic peptides derived from MBP, the measurement of anti-idiotypic antibody and the detection of MBP in human serum.

From an examination of electroimmunoblots and peptide maps, a mouse monoclonal antibody to human myelin basic protein MBP was shown to react with the amino acid sequence Ala-Ser-Asp-Tyr-Lys-Ser which is located in the C-terminal half of MBP. Although a completely different immunization schedule was used by Sires et al. (1981) they obtained a monoclonal antibody reacting with the same determinant. In contrast to results with other monoclonal antibodies to globular proteins (Todd et al. 1982) this monoclonal antibody seems to react with a sequential rather than a topographical determinant.

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.