Fusion expression of human X-box binding protein 1 and preparation of polyclonal antibody against protein

AIM:To express prokaryotically and purify the unsplicedhuman X-box binding protein1(XBP1u)recombinant plasmid and prepare polyclonal antibody against protein.METHORDS:We transformed the recombinant plasmid pET32a-XBP1u into host bacterium E.coli BL21(DE3),induced its expressionwith IPTG,and purified the protein with Ni2+-NTA collumn after identified by SDS-PAGE,then immunized two New Zealand rabbits with the purified protein as antigen after renaturation with dialytic bag,then collected blood from the heart after the last immunization,detected the titer of antiserum by ELISA,detected the antiserum specificity by Western Blot and immunohistochemistry.RESULTS:XBP1u expressed in the inclusion body of E.coli BL21,the titer of antiserum was beyond 1∶64 000 after immunizerabbit with purification and renaturation XBP1u protein.Besides,it is confirmed that the antiserum reacted specifically to the XBP1u protein which is Mr 33×103 by Western Blot.It is successful to detect the expression in hepatocyte with immunohistochemistry.CONCLUSION:It is successful to induce XBP1u expressionand purification.The highly specific and titer polyclonal antibody have been obtained on basis of this purified protein.