【摘要】 为制备兔抗猪CD163多克隆抗体,采用RT-PCR方法从猪PAM RNA中扩增CD163胞质外区部分基因SRCR4-5,克隆至pGEX-6p-1原核表达载体,在IPTG诱导下获得约50 000以包涵体形式表达的重组蛋白。WesternBlotting分析表明,表达产物能够被GST单抗所识别。用该重组蛋白免疫兔制备多抗血清,经间接ELISA检测,多抗血清效价可达105,流式检测发现可与Marc-145和PAM上的天然CD163分子相互作用。抗体阻断试验证明制备的多抗血清可以部分阻断PRRSV感染。本试验为进一步研究PRRSV与受体-CD163相互作用机制打下了基础。更多还原

【Abstract】 The extracellular region of porcine CD163 gene,SRCR4-5,was amplified from total RNA of porcine alveolar macrophages and subcloned into prokaryotic expression vector pGEX-6p-1.Followed by inducing with IPTG,a 50 000 fusion protein which could be recognized by anti-GST monoclonal antibody was expressed.After purified,the fusion protein was employed to immunize rabbits to generate rabbit anti-CD163 polyclonal antibodies.The polyclonal antibodies had a titer of 105 in GST-SRCR4-5 by indirect ELISA,and it also could partly block the infection of PRRSV to porcine alveolar macrophages in antibody-blocking assay.The results suggested that the polyclonal antibody was of biological function,and could be employed in the further studies on CD163 receptor.更多还原