Preparation and functional identification of a monoclonal antibody against the recombinant soluble human NKp30 receptor

NKp30 is an important activating receptor of human natural killer (NK) cells that participates in NK cell activation and cytotoxicity against tumor and infected cells. To study the function of NKp30, anti-human NKp30 monoclonal antibody was prepared. The human NKp30 ectodomain (rhNKp30) was expressed in Escherichia coli as inclusion bodies and refolded using the dilution method. The refolded rhNKp30 was purified by immobilized metal affinity chromatography. The activity of soluble rhNKp30 was confirmed by flow cytometry and NK cytotoxicity assays. Four hybridoma cell lines producing monoclonal antibodies against rhNKp30 were obtained. One of the monoclonal antibodies, designated as “3G5”, was highly specific and could be used in western blotting, immunoprecipitation, ELISA, and flow cytometry assays. The preparation of soluble rhNKp30 and a monoclonal antibody against NKp30 may provide useful tools for further functional studies of human NKp30.