Prokaryotic expression of the extracellular region of human CD1d and preparation of its polyclonal antibody

AIM: To explore the prokaryotic expression of the extracellular region of human CD1d (hCD1d) and prepare its polyclonal antibody. METHODS: The gene encoding the extracellular region of hCD1d was amplified by PCR and cloned into prokaryotic expression vector pET28, then expressed in E.coli BL21(DE3) with IPTG induction. The recombinant protein was purified by Ni2+-NTA agarose column and then used as immunogen to immunize the mouse. The generated polyclonal antibody was evaluated by ELISA, Western blot and immunohistochemical staining (IHC), respectively. RESULTS: The recombinant extracellular region of hCD1d was successfully expressed and purified. The polyclonal antibody with high titer and high specificity was obtained, which could recognize the native hCD1d in the human small intestinal tissues. CONCLUSION: The recombinant extracellular region of hCD1d has been obtained. The antibody with high titer and high specificity against the extracellular region of hCD1d from the mouse has been successfully prepared, which lays a foundation for further research into the detection and functional study of CD1d.