Specificity and VH sequence of two monoclonal antibodies against the N-terminus of dystrophin.
We have used a random library of 15-mer peptides expressed on phage to show that two monoclonal antibodies (mAbs) require only the first three amino acids of dystrophin (Leu-Trp-Trp) for binding. Since the mAbs recognize dystrophin in frozen muscle sections, the results suggest that this hydrophobic N-terminus of dystrophin is accessible to antibody in situ. Quantitative binding studies suggested minor differences in specificity between the two mAbs, so the Ig heavy-chain variable region (VH) sequences of the two hybridomas were determined by RT-PCR and cDNA sequencing. After elimination of PCR errors, the two cDNA sequences were found to be identical except for five somatic mutations which resulted in three amino acid changes in the second hypervariable region (CDR2). The results suggest that the two hybridomas originated from the same lymphocyte clone in a germinal centre of the spleen, but underwent different point mutations and subtype switches during clonal expansion to form blast cells.
Morris
G E
GE
MRIC Biotechnology Group, North East Wales Institute, Deeside, Clwyd, U.K.
Nguyen
C
C
Nguyen thi Man
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D001483:Base Sequence; D002460:Cell Line; D018076:DNA, Complementary; D016189:Dystrophin / Q000737:chemistry / Q000276:immunology*; D007143:Immunoglobulin Heavy Chains / Q000276:immunology*; D007135:Immunoglobulin Variable Region / Q000276:immunology*; D051379:Mice; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid
D000911:Antibodies, Monoclonal; D018076:DNA, Complementary; D016189:Dystrophin; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region