Preparation and functional characterization of the monoclonal antibody HAb18Gedomab1

AIM: To obtain mouse anti-human monoclonal antibodies against recombinant extracellular domain of HAb18G (HAb18Ged), and to analyze and identify its character and biological function. METHODS: Balb/c mice were immunized with HAb18Ged. Hybridoma cell was screened by cell fusion and subcloning approach. The monoclonal antibody in the ascites was purified by ion exchange chromatography and was identified by fluorescence-activated cell sorting analysis (FACs) and immunohistochemistry. Gelatin zymography and collagenase type I zymography were used to analyze the effects of HAb18Gedomab1 on activation and production of matrix metalloproteinase (MMPs); Matrigel-boyden degradation chamber method was used to evaluate the infiltrative cells ratio. RESULTS: A hybridoma cell HAb18Gedomab1 stably secreting anti-HAb18Ged monoclonal antibody was obtained. The titer of this McAb in ascites was 1:10⁶. The purity of the McAb was higher than 90%. The McAb belonged to IgG₁ subclass. HAb18Gedomab1 showed high specificity and affinity to the antigen of FHCC-98 cell membrane and the tissue of hepatocellular carcinoma. The McAb induced production and activation of MMP-2, MMP-9, MMP-1 and MMP-8 in mouse fibroblast cells (3T3), and also promoted the degradation of reconstituted basement membrane. CONCLUSION: HAb18Gedomab1 can bind specifically to HAb18Ged protein. The McAb can also induce production and activation of MMPs and promote the degradation of reconstituted basement membrane.