Kuroki M, Kuroki M, Koga Y, Matsuoka Y
J Immunol. 1984 Oct;133(4):2090-7.
Eleven hybridoma clones producing monoclonal antibodies to carcinoembryonic antigen (CEA) were prepared by fusions of the mouse myeloma cell line P3-X63-Ag8-U1 with splenocytes of BALB/c mice immunized with highly purified CEA. Their specificities were systematically analyzed by radioimmunoassay for reactivity with CEA and four related normal antigens: nonspecific cross-reacting antigen (NCA), nonspecific cross-reacting antigen-2 (NCA-2), normal fecal antigen-1 (NFA-1), and normal fecal antigen-2 (NFA-2). Antibody-antigen profiles revealed four groups of hybridomas. The Group I antibodies from two clones reacted with sites shared among CEA, NCA, NCA-2, and NFA-2. The Group II antibody from one clone recognized an epitope common to CEA, NCA-2, and NFA-2. The Group III antibodies from five clones reacted with sites commonly residing on CEA, NCA-2, NFA-1, and NFA-2, whereas antibodies of Group IV from three clones bound only CEA. Mutual inhibition assays between antibodies in the respective groups for CEA binding further revealed that at least eight different epitopes can be identified on the CEA molecule: two in Group I, one in Group II, three in Group III, and two in Group IV, respectively. Three monoclonal antibodies in Group IV appeared to be specific for CEA, but they did not react with any of five purified CEA preparations other than that used for immunization. In 13 sera from 50 various cancer patients with elevated CEA levels, however, we detected the CEA molecule reactive with these antibodies, indicating that the epitopes recognized by the Group IV antibodies may be of allotypic characters. These epitopes were unrelated to main blood group antigens and did not show any organ specificity. All of the eight epitopes identified in this study were found to be resistant to neuraminidase, mixed glycosidases, and Smith degradation (SI-stage) as determined by a competitive radioimmunoassay, suggesting that they reside on the protein moiety of the CEA molecule. Six epitopes were sensitive to reduction and alkylation, but two other epitopes (one in Group II and the other in Group III) were resistant to these treatments. This result indicates that the latter two epitopes are conformation-independent.
通过将小鼠骨髓瘤细胞系P3-X63-Ag8-U1与用高度纯化的癌胚抗原（CEA）免疫的BALB/c小鼠的脾细胞融合，制备了11个产生抗癌胚抗原单克隆抗体的杂交瘤克隆。通过放射免疫分析系统地分析了它们与CEA及四种相关正常抗原的反应性，这四种正常抗原分别为：非特异性交叉反应抗原（NCA）、非特异性交叉反应抗原-2（NCA-2）、正常粪便抗原-1（NFA-1）和正常粪便抗原-2（NFA-2）。抗体-抗原谱显示有四组杂交瘤。来自两个克隆的I组抗体与CEA、NCA、NCA-2和NFA-2共有的位点发生反应。来自一个克隆的II组抗体识别CEA、NCA-2和NFA-2共有的一个表位。来自五个克隆的III组抗体与CEA、NCA-2、NFA-1和NFA-2上共有的位点发生反应，而来自三个克隆的IV组抗体仅与CEA结合。各小组抗体之间针对CEA结合的相互抑制试验进一步表明，在CEA分子上可鉴定出至少八个不同的表位：I组两个，II组一个，III组三个，IV组两个。IV组中的三种单克隆抗体似乎对CEA具有特异性，但它们除了与用于免疫的CEA制剂外，不与其他五种纯化的CEA制剂发生反应。然而，在50例CEA水平升高的各种癌症患者的13份血清中，我们检测到了与这些抗体发生反应的CEA分子，这表明IV组抗体识别的表位可能具有同种异型特征。这些表位与主要血型抗原无关，也没有显示出任何器官特异性。通过竞争性放射免疫分析确定，本研究中鉴定出的所有八个表位均对神经氨酸酶、混合糖苷酶和史密斯降解（SI阶段）具有抗性，这表明它们位于CEA分子的蛋白质部分。六个表位对还原和烷基化敏感，但另外两个表位（一个在II组，另一个在III组）对这些处理具有抗性。这一结果表明后两个表位与构象无关。
