Preparation and identification of the monoclonal antibody against FcεRIα

Objective To prepare and initially identify the monoclonal antibody against FceRIα, so as to provide basis for further reseach on anaphylactic diseases.Methods BALB/C mice was immunized with the recombination protein,and the hybridoma cell line which stably secreted FceRIαmAb was prepared via fusion,screening and cloning.Then ElISA,immunofluorescence labeling method, fluorescent microscopy examination,flow cytometry and Western-Blot assay were adopted to perform initial identification of the monoclonal antibody.Results Two cell strains were screened which stably secreted FceRIαmAb against FceRIα.Flow cytometry showed that the binding rate of the antibodies to CH03D10 cells was over 95%.The antibodies named as B4C2 and B5B4 displayed strong specifility and high affinity.The titration of antibodies in supernatant and mouse ascites were higher than 105. The immunofluorescence assay demonstrated that the antibodies specifically binded with CH03D10 cells,and Western Blot showed the light and heavy chains of the antibodies were 28 Kd and 50 Kd respectively. Conclusion The prepared two strains of specific mAbs against human FceRIαcan bind with FceRI on cell surface,which can be used as a tool of further research on mast cell function and clinical therapy of allergosis.