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目的:制备和鉴定抗人肺鳞癌单克隆抗体(mAb),为肺癌的早期诊断和靶向治疗提供候选抗体药物及为肺癌抗原的筛选提供工具。方法:用肺癌细胞株YTMLC-90做抗原免疫BALB/c小鼠,取其脾细胞与小鼠骨髓瘤细胞Sp2/0融合,间接ELISA和有限稀释法筛选抗体阳性的杂交瘤细胞获得mAb。染色体计数法鉴定杂交瘤细胞,并进行mAb同种型鉴定;对细胞培养上清液和腹水中的mAb采用间接ELISA法进行初步特异性检测及效价测定;采用Western blot法鉴定其与不同组织抗原结合活性;采用免疫组化染色法鉴定其在肺癌组织中的分布。结果:筛选出5株可稳定分泌抗肺鳞癌mAb的杂交瘤细胞株E2、F9、E11、D5和C6,分泌的抗体均为IgG1亚类,κ亚型。C6染色体数目为89～112条,细胞培养上清和腹水效价分别为1∶1 004和1∶104,ELSA和Western blot结果显示其能特异性与肺癌细胞结合,未见与其他高发癌组织和正常组织结合。结论:成功地制备了能够特异识别肺癌组织的mAb。
OBJECTIVE: To prepare and identify anti-human lung squamous cell carcinoma monoclonal antibody (mAb), to provide candidate antibody drugs for the early diagnosis and targeted therapy of lung cancer and to provide a tool for the screening of lung cancer antigens. Methods: BALB / c mice were immunized with lung cancer cell line YTMLC-90. Sp2 cells were fused with Sp2 / 0 of mouse myeloma cells. The mAbs were screened by indirect ELISA and limited dilution method. The hybridoma cells were identified by chromosome counting and the mAb isotype was identified. The mAbs in the cell culture supernatants and ascites were detected by indirect ELISA and the titer was determined. Western blot was used to identify the hybridomas with different tissues Antigen binding activity; Immunohistochemical staining to identify its distribution in lung cancer. Results: Five strains of hybridoma cell lines E2, F9, E11, D5 and C6 that could stably secrete anti-squamous cell carcinoma mAb were screened. The secreted antibodies were all IgG1 subclass and κ subtype. The numbers of C6 chromosomes ranged from 89 to 112. The cell culture supernatants and ascites titers were 1: 1 004 and 1: 104, respectively. ELSA and Western blot showed that they could specifically bind with lung cancer cells. Normal tissue union. Conclusion: mAbs that specifically recognize lung cancer tissue were successfully prepared.