Dai Hua, Chen Junhua, Zhang Haixia, Shen Ying, Chen Xiang, Jiao Xin'an
Jiangsu Key Laboratory of Zoonosis, Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Medical School, Yangzhou University, Yangzhou 225001, China.
Jiangsu Key Laboratory of Zoonosis, Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Medical School, Yangzhou University, Yangzhou 225001, China. *Corresponding author, E-mail:
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2017 Jul;33(7):986-990.
Objective To prepare and characterize the monoclonal antibodies (mAbs) against chicken interleuikin-4 (ChIL-4). Methods Using lymphocyte hybridoma technique, 8-week-old BALB/c mice were immunized with purified recombinant ChIL-4 (rChIL-4) with GST tag (GST-ChIL-4), and rChIL-4 with histidine tag (His-ChIL-4) was utilized to set up an indirect ELISA to screen positive hybridomas. Limited dilution methods were used to generate hybridoma cell lines stably secreting mAbs against ChIL-4. The specificity of the mAbs was identified by indirect ELISA, immunofluorescent assay (IFA) and Western blot. Results Four mAbs against rChIL-4 were obtained and named 6A8, 18F12, 19F1 and 20G3. ELISA titers of the 4 mAbs were (1.6~6.4)×10. ELISA, IFA and Western blot showed that the 4 mAbs could only react with rChIL-4, but not with the other proteins like bovine IL-4 (BovIL-4), chicken IFN-γ (ChIFN-γ). Conclusion The mAbs against rChIL-4 were successfully obtained.
目的 制备并鉴定抗鸡白细胞介素-4（ChIL-4）的单克隆抗体（mAb）。方法 采用淋巴细胞杂交瘤技术，用带谷胱甘肽S-转移酶标签（GST）的纯化重组ChIL-4（rChIL-4）免疫8周龄BALB/c小鼠，并用带组氨酸标签（His）的rChIL-4建立间接ELISA筛选阳性杂交瘤。采用有限稀释法制备稳定分泌抗ChIL-4 mAb的杂交瘤细胞系。通过间接ELISA、免疫荧光分析（IFA）和蛋白质印迹法鉴定mAb的特异性。结果 获得4株抗rChIL-4的mAb，分别命名为6A8、18F12、19F1和20G3。这4株mAb的ELISA效价为（1.6～6.4）×10。ELISA、IFA和蛋白质印迹结果显示，这4株mAb仅与rChIL-4反应，不与牛白细胞介素-4（BovIL-4）、鸡干扰素-γ（ChIFN-γ）等其他蛋白质反应。结论 成功获得了抗rChIL-4的mAb。
