Direct determination of molecular ratios of peptides coupled via N-succinimidyl 3-(2-pyridyldithio)propionate to carrier proteins using high performance liquid chromatography.
A simple, rapid and reproducible method is presented for direct determination of the substitution ratio of a carrier protein with a synthetic nonradioactively labeled peptide. The peptide was covalently linked by a thiol group of a cysteine residue to the immunogenic carrier protein using the heterobifunctional reagent N-succinimidyl 3-(2-pyridyldithio)propionate. The substitution ratio was determined after reductive cleavage of the intermolecular disulfide bond between peptide and carrier and the amount of carrier and peptide quantitated directly by calibrated HPLC analysis within 15 min.
Bläsi
U
U
Institute of Genetics and Microbiology, University of Munich, F.R.G.
Linke
R P
RP
Lubitz
W
W
D000818:Animals; D000917:Antibody Formation; D001426:Bacterial Proteins / Q000008:administration & dosage / Q000276:immunology; D002352:Carrier Proteins / Q000008:administration & dosage / Q000276:immunology; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003432:Cross-Linking Reagents; D010455:Peptides / Q000008:administration & dosage / Q000276:immunology; D011487:Protein Conformation; D011817:Rabbits; D013388:Succinimides; D013954:Thyroglobulin / Q000008:administration & dosage / Q000276:immunology; D014764:Viral Proteins
D001426:Bacterial Proteins; D002352:Carrier Proteins; D003432:Cross-Linking Reagents; C029555:E protein, bacteriophage X174; D010455:Peptides; D013388:Succinimides; D014764:Viral Proteins; C018151:N-succinimidyl 3-(2-pyridyldithio)propionate; D013954:Thyroglobulin