AIM: To prepare the monoclonal antibody (mAb) against human growth differentiation factor 15 (GDF15). METHODS: The expression vector pGEX-4T-2-gdf15 was constructed and transformed into E.coli Top10F' for expression. Then the purified fusion protein was used to immunize the BALB/c mice. The mouse myeloma cells (Sp2/0) were fused with spleen cells from the BALB/c immunized by the purified protein. Subsequently, limited dilution method was used three times to screen hybridoma cell lines. The titer of the mAb was detected by ELISA and its specificity was analyzed by Western blot. The serum level of GDF15 in hepatocellular carcinoma (HCC) and health people was measured by co-immunoprecipitation (IP) method. RESULTS: The GST-GDF15 fusion protein was successfully expressed and purified. One hybridoma cell line designated 9G3 against GDF15 was obtained. IP and mass spectrometric analysis revealed that the mAb recognized GDF15 in human sera with high specificity. The level of GDF15 in HCC patients was much higher than that in health people. CONCLUSION: The success in mouse anti-GDF15 mAb preparation provides the basis for further developing HCC diagnose kit.
AIM: To prepare the monoclonal antibody (mAb) against humangrowth differentiation factor 15 (GDF15). METHODS: The expression vector pGEX-4T-2-gdf15 was constructed and transformed into E.coli Top10F' for expression. Then the purified fusion protein was used to immunize the BALB/c mice. The mousemyeloma cells (Sp2/0) were fused with spleen cells from the BALB/c immunized by the purified protein. Subsequently, limited dilution method was used three times to screen hybridoma cell lines. The titer of the mAb was detected by ELISA and its specificity was analyzed by Western blot. The serum level of GDF15 in hepatocellular carcinoma (HCC) and health people was measured by co-immunoprecipitation (IP) method. RESULTS: The GST-GDF15 fusion protein was successfully expressed and purified. One hybridoma cell line designated 9G3 against GDF15 was obtained. IP and mass spectrometric analysis revealed that the mAb recognized GDF15 in human sera with high specificity. The level of GDF15 in HCC patients was much higher than that in health people. CONCLUSION: The success in mouse anti-GDF15 mAb preparation provides the basis for further developing HCC diagnose kit.
Authors: Nisha Bansal; Leila Zelnick; Michael G Shlipak; Amanda Anderson; Robert Christenson; Rajat Deo; Christopher deFilippi; Harold Feldman; James Lash; Jiang He; John Kusek; Bonnie Ky; Stephen Seliger; Elsayed Z Soliman; Alan S Go Journal: Clin Chem Date: 2019-10-02 Impact factor: 8.327