Johansson A, Sandström P, Ullén A, Behravan G, Erlandsson A, Levi M, Sundström B, Stigbrand T
Department of Immunology, Umeå University, Sweden.
Cancer Res. 1999 Jan 1;59(1):48-51.
Due to their abundance in epithelial cells and deposition in necrotic regions intratumorally, cytokeratins (CKs) have been established as valuable targets for both radioimmunolocalization and radioimmunotherapy. The target epitope for the monoclonal anti-CK8 antibody, TS1, 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 identified. The epitope retains its helical structure, as shown with circular dichroism spectroscopy, although the length of the peptide (ie., >20 aa) is crucial for maintenance of immunoreactivity. To determine which aa residues are crucial for binding to the monoclonal antibody, alanine scanning was performed on a 26-mer covering aa 340-365, with the sequence QRGELAIKDANAKLSELEAALQRAKQ. The 26 modified peptides were evaluated using ELISA and BIAcore technology. The uniqueness of this epitope has been established by data base sequence comparisons.
由于细胞角蛋白（CKs）在上皮细胞中含量丰富且在肿瘤内坏死区域沉积，它们已成为放射免疫定位和放射免疫治疗的重要靶点。用于实验性放射免疫定位和放射免疫治疗的单克隆抗CK8抗体TS1的靶表位，是通过合成覆盖整个CK8分子的96个重叠肽段来确定的。确定了一个高度保守的肽序列，跨越氨基酸（aa）343 - 357，覆盖螺旋2B结构域中的不连续表位。尽管肽的长度（即>20个氨基酸）对于维持免疫反应性至关重要，但如圆二色光谱所示，该表位保留其螺旋结构。为了确定哪些氨基酸残基对于与单克隆抗体结合至关重要，对覆盖aa 340 - 365的26聚体（序列为QRGELAIKDANAKLSELEAALQRAKQ）进行了丙氨酸扫描。使用酶联免疫吸附测定（ELISA）和生物传感器技术（BIAcore）对26个修饰肽进行了评估。通过数据库序列比较确定了该表位的独特性。
