重组鸡白细胞介素2的诱导表达与生物学活性测定

 将去除信号肽的鸡IL-2基因编码框克隆到原核表达载体pBAD/His B，实现了重组鸡IL-2(rchIL-2)蛋白在大肠杆菌中的高效表达。SDS-PAGE分析显示，表达蛋白的分子量约为18 kD。用rchIL-2为抗原制备了单克隆抗体和多克隆抗体，建立了检测rchIL-2含量的抗原捕获ELISA，探讨了天然chIL-2蛋白的体外表达动力学。非变性条件下纯化的rchIL-2 (0.4 ng) 对Con A活化的鸡T淋巴细胞具有明显的增殖活性，而对鸭及鹅的T淋巴细胞无增殖活性。抗chIL-2多克隆抗体可完全中和rchIL-2和天然chIL-2蛋白的生物学活性，而单克隆抗体无中和rchIL-2和天然chIL-2蛋白生物学活性的功能。这些研究结果证实，大肠杆菌表达的rchIL-2及其多克隆抗体拥有生物学功能，而获得的单克隆抗体不具有chIL-2的中和特性。

In vitro Expression and Bioactivity of Chicken Interleukin-2

In this study, the recombinant chicken interleukin-2 (rchIL-2) was successfully produced in the pBAD/His B expression system induced by L-(+)-arabinose. By SDS-PAGE analysis, the observed molecular weight of rchIL-2 fusion protein without the signal peptide was 18 kD. Monoclonal and polyclonal antibodies specific for rchIL-2 were produced. These antibodies were used to develop a mAb-based antigen capture ELISA (AC-ELISA). Based on the constructed AC-ELISA, in vitro kinetics of endogenous chicken IL-2 production were detected. The soluble rchIL-2 protein purified under native conditions was shown to activate the proliferation of chicken lymphocytes stimulated by Con A for 24 h in a dose-dependent manner. This activity cannot be neutralized by mAbs to chIL-2, but can be neutralized by anti-chIL-2 polyclonal antibody. These results demonstrated that the E.coli-derived rchIL-2 and polyclonal antibody to chIL-2 have bioactivity.