CAJ | 학술논문

目的：利用原核表达系统表达人肝脏凝血因子Ⅺ（ coagulation factor Ⅺ， F11）蛋白，并制备F11的兔多克隆抗体。方法从cDNA文库中筛选并扩增F11基因，将其分别与带有HIS标签的pET-32a及GST标签pGEX-4 T-1载体连接，获得表达质粒，将表达质粒转入BL21菌中表达F11融合蛋白（分别含有HIS、GST-Tag），通过SDS-PAGE进行鉴定。获得的F11融合蛋白经纯化后作为免疫原及检测原，制备兔多克隆抗体，Western blot分析获得的兔多克隆抗体的特异性。结果成功构建pET-32a-F11、pGEX-4t-1-F11表达质粒，获得两种标签的融合蛋白。将重组蛋白纯化后作为免疫原免疫兔，取得了能与重组蛋白特异性结合的兔血清。结论成功制备了F11重组蛋白及F11兔多克隆抗体，为进一步研发F11诊断试剂打下了基础。
목적：이용원핵표체계통표체인간장응혈인자Ⅺ（ coagulation factor Ⅺ， F11）단백，병제비F11적토다극륭항체。방법종cDNA문고중사선병확증F11기인，장기분별여대유HIS표첨적pET-32a급GST표첨pGEX-4 T-1재체련접，획득표체질립，장표체질립전입BL21균중표체F11융합단백（분별함유HIS、GST-Tag），통과SDS-PAGE진행감정。획득적F11융합단백경순화후작위면역원급검측원，제비토다극륭항체，Western blot분석획득적토다극륭항체적특이성。결과성공구건pET-32a-F11、pGEX-4t-1-F11표체질립，획득량충표첨적융합단백。장중조단백순화후작위면역원면역토，취득료능여중조단백특이성결합적토혈청。결론성공제비료F11중조단백급F11토다극륭항체，위진일보연발F11진단시제타하료기출。
Objective To express recombinant coagulation factor Ⅺ( F11 ) using prokaryotic expression system , and to prepare rabbit polyclonal antibodies against F 11 .Methods F11 gene was obtained from the cDNA library , and connected with pET-32a and pGEX-4T-1 vector to get the expression plasmid respectively .F11 fusion protein was expressed by putting the ex-pression plasmid into BL21, and identified by SDS-PAGE.Rabbits were immunited with the purified F11 fusion protein for the preparation of polyclonal antibodies .The specificity of rabbit polyclonal antibodies was identified by Western blot .Resulst Recombinant plasmids pET-32a-F11 and pGEX-4T-1-F11 were successfully constructed .And F11 rabbit serum identified corresponding recombinant proteins after the purified fusion protein as immunogen to immune rabbit.Conclusion Rabbit polyclonal antibodies against recombinant F 11 protein are pre-pared successfully , which lays a foundation for further development of F 11 diagnostic reagents .