[Preparation and characterization of monoclonal antibody against HCCR protein].

Aim: Preparation of monoclonal antibody (mAb) to HCCR, which is a candidate biomarker for human hepatocellular carcinoma (HCC).

Methods: The recombinant protein HCCR-1(167-360); was expressed and was used as immunogen to immunize mouse for generation of mAb against HCCR. The protein Ep-HCCR, which displayed a epitope of HCCR, was also expressed and purified to use to detect serum antibody titer and to screen the positive clones of hybridmas. The properties of HCCR antibody were analyzed by ELISA, Western blot, immunofluorescence and immunohistochemistry.

Results: A hybridmas clone, which secreted anti-HCCR mAb, was obtained. The affinity constant (Kaff) of the mAb is 5.4×10(6); L/mol analyzed by ELISA; Western blot showed that the mAb could specifically recognize HCCR-1 and HCCR-2 expressed in HepG2 cells; The mAb was also used to detect the expression of HCCR proteins in hepatoma cells and HCC tissues. The results of immunofluorescence indicated that HCCR proteins mainly localized on the plasma membrane and cytoplasm of HepG2 cells. In addition, HCCR was found high-expressed in HCC tissues but not in normal liver tissue detected by Immunohistochemistry.

Conclusion: A specific mAb against HCCR was successfully generated, which laid the foundation for establishing HCC detection method based on HCCR.