Epitope specificity of the monoclonal anticytokeratin antibody TS1

Due to their abundance in epithelial cells and deposition in necrotic regio ns intratumorally, cytokeratins (CKs) have been established as valuable tar gets for both radioimmunolocalization and radioimmunotherapy. The target ep itope for the monoclonal anti-CK8 antibody, TSI, used for both experimental radioimmunolocalization and radioimmunotherapy, was determined by means of synthesis of 96 overlapping peptides that covered the entire CK8 molecule. A highly conserved peptide sequence, spanning amino acids (aa) 343-357 and covering the discontinuous epitope in the helical 2B domain, was identifie d. The epitope retains its helical structure, as shown with circular dichro ism spectroscopy, although the length of the peptide (i.e., >20 aa) is cruc ial for maintenance of immunoreactivity, To determine which aa residues are crucial for binding to the monoclonal an tibody, alanine scanning was performed on a 26-mer covering aa 340-365, wit h the sequence QRGELAIKDANAKLSELEAALQRAKQ. The 26 modified peptides were ev aluated using ELISA and BIAcore technology. The uniqueness of this epitope has been established by data base sequence comparisons.