Synthetic peptides representing different areas of the CEA molecule were used as immunogens for the development of anti-CEA antibodies. Both polyclonal and monoclonal antibodies were generated using peptides composed of CEA amino acid positions 99–128 and 585–613, respectively. One MAb, designated CP4, generated using the CEA peptide 99–128, was chosen for a more detailed analysis of reactivity. MAb CP4 reacts in solid phase RIAs with CEA peptide 99–128 immunogen and purified native CEA. CP4 did not react with purified non- specific cross reacting antigen (NCA), even though there were two single amino acid differences between NCA and CEA in the 29 amino acid peptide. The affinity constants of CP4 for the CEA peptide 99–128 and native CEA are 4.07 × 10⁹ M⁻¹ and 5.75 × 10⁸ M⁻¹, respectively. When CP4 was reacted with purified CEA in Western blotting experiments, the Mr 180,000 glycoprotein characteristic of CEA was detected, but CP4 reacted to various size entities in tumor cell extracts. The results of liquid competition RIAs showed that the epitope that MAb CP4 recognized on native CEA is not available for binding when CEA is in solution. Physical (adsorption to a solid matrix) or chemical (deglycosylation or formalin-fixation) alteration of CEA is required for binding of CP4 to CEA. MAb CP4 reacted approximately 1,000-fold greater to deglycosylated CEA than native CEA. Immunohistochemical studies using formalin-fixed paraffin-embedded tissue sections demonstrated that, among carcinomas, CP4 reacts selectively with colorectal carcinomas, while normal colon is negative. Although stomach carcinoma is negative, dysplastic lesions and areas of intestinal metaplasia are reactive. Two of 7 normal stomach tissues showed focal cytoplasmic reactivity of the surface epithelium. CP4, therefore, appears to react with an epitope with highly restricted expression in colorectal carcinoma. These studies demonstrate the complexities in dealing with an anti-peptide MAb with reactivity to an epitope which is accessible only under certain conditions.

代表CEA分子不同区域的合成肽被用作产生抗CEA抗体的免疫原。多克隆抗体和单克隆抗体都是使用分别由CEA氨基酸99-128和585-613位氨基酸组成的肽生成的。选择一种使用CEA肽99-128生成的称为CP4的单克隆抗体，以进行更详细的反应性分析。MAb CP4在固相RIA中与CEA肽99-128免疫原和纯化的天然CEA反应。即使在29个氨基酸的肽段中，NCA和CEA之间存在两个单氨基酸差异，CP4也不会与纯化的非特异性交叉反应抗原（NCA）发生反应。CP4对CEA肽99-128和天然CEA的亲和常数为4.07×10 ⁹ M ^-1和5.75×10 ⁸M ^-1， 分别。在Western印迹实验中，当CP4与纯化的CEA反应时，检测到CEA的180,000糖蛋白先生特征，但CP4与肿瘤细胞提取物中的各种大小的实体发生了反应。液体竞争RIA的结果表明，当CEA在溶液中时，MAb CP4在天然CEA上识别的表位不可用于结合。CP4与CEA的结合需要对CEA进行物理（吸附到固体基质）或化学（去糖基化或福尔马林固定）的改变。与天然CEA相比，MAb CP4对去糖基化CEA的反应大约高1000倍。使用福尔马林固定石蜡包埋的组织切片进行的免疫组织化学研究表明，在癌症中，CP4与结直肠癌选择性反应，而正常结肠为阴性。尽管胃癌是阴性的 增生性病变和肠上皮化生区域是反应性的。7个正常胃组织中有2个显示了表面上皮的局部胞浆反应性。因此，CP4似乎与在结肠直肠癌中表达高度受限的表位反应。这些研究表明，处理与表位具有反应性的抗肽单克隆抗体的复杂性仅在某些条件下才可达到。