Affinity purification of two populations of antibodies against format determinants of synthetic myelin basic protein peptide S82 from S82-AH- and S82-CH-Sepharose 4B columns.
Two different kinds of immunosorbents were prepared that contained the synthetic myelin basic protein didecapeptide S82 (TTHYGSLPQKAQGHRDQDEG)--one coupled with AH-Sepharose 4B through hexanoate spacers to the C-terminal glycyl residue; the other, with CH-Sepharose 4B through hexanoate spacers to the N-terminal threonine residue. An antiserum rich in antibodies to a format determinant of S82 was passed through each column, and, by means of affinity purification, two homogeneous populations of anti-format antibodies were obtained, each with a binding affinity of 1 X 10(8)M-1 for S82. The population recovered from S82-AH-Sepharose 4B cross-reacted to a considerable extent with synthetic peptide S8 (GSLPQKAQGHRPQDENG) but only to a limited extent with S79 (AQGHRPQDEG). The population recovered from S82-CH-Sepharose 4B cross-reacted poorly, if at all, with S8. An equimoler mixture of S8 + S79, however, reacted well with either population of anti-format antibodies, thus showing that the mixture could mimic the format of S82. It was concluded that secondary structural conformation of S82 could be preserved during the coupling procedure and that the resulting immunosorbents could be used for the affinity purification of anti-S82 antibodies to the format determinants.
D000818:Animals; D000906:Antibodies / Q000302:isolation & purification*; D002846:Chromatography, Affinity; D000939:Epitopes / Q000276:immunology; D004676:Myelin Basic Protein / Q000276:immunology*; D010446:Peptide Fragments / Q000276:immunology*; D011817:Rabbits / Q000276:immunology
D000906:Antibodies; D000939:Epitopes; D004676:Myelin Basic Protein; D010446:Peptide Fragments