CAJ | 학술논문

试验旨在制备鼠抗鸡Toll样受体2(chTLR2)的多克隆抗体。采用RT-PCR方法从活化的鸡外周血淋巴细胞中扩增其胞外区片段chTLR2(chTLR2t1和chTLR2t2),并构建这两个片段的原核表达重组质粒,优化蛋白质的表达条件,SDS-PAGE和Western-blot法检测融合蛋白,将融合蛋白作为免疫原免疫Balb/c小鼠,利用相应的原核表达的全长蛋白检测特异性抗体滴度。结果显示,成功获得chTLR2的胞外区片段,构建的原核表达重组质粒经IPTG诱导能成功表达预期的重组蛋白,免疫小鼠可产生特异性抗体,抗体效价分别为1：16000和1：10000。表明制备的鼠抗chTLR2多抗具有良好的特异性,为进一步研究chTLR2的功能提供了重要材料。
시험지재제비서항계Toll양수체2(chTLR2)적다극륭항체。채용RT-PCR방법종활화적계외주혈림파세포중확증기포외구편단chTLR2(chTLR2t1화chTLR2t2),병구건저량개편단적원핵표체중조질립,우화단백질적표체조건,SDS-PAGE화Western-blot법검측융합단백,장융합단백작위면역원면역Balb/c소서,이용상응적원핵표체적전장단백검측특이성항체적도。결과현시,성공획득chTLR2적포외구편단,구건적원핵표체중조질립경IPTG유도능성공표체예기적중조단백,면역소서가산생특이성항체,항체효개분별위1：16000화1：10000。표명제비적서항chTLR2다항구유량호적특이성,위진일보연구chTLR2적공능제공료중요재료。
To prepare mouse anti-chicken Toll-like receptor 2 (chTLR2) polyclonal antibodies, four-week-old spe-cific-pathogen-free ( SPF) chickens were infected with Salmonella enteritidis via oral route. Two fragments of extracellular region of chTLR2 genes (chTLR2t1 and chTLR2t2) from infected chicken peripheral blood lymphocytes were by RT-PCR, and the two prokaryotic expression vectors of chTLR2 were constructed for protein expression. The two fusion proteins were detected by SDS-PAGE and Western-blot assay, and were used to immunize eight-week-old Balb/c mice to prepare poly-clonal antibodies. The extracellular region of chTLR2 was successfully expressed in prokaryotic expression vector. The two recombinant proteins induced by IPTG produced the antibodies against mice with the antibodies titer being 1 : 16 000 and 1 : 10 000, respectively, suggesting a good specificity of the polyclonal antibodies.