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Screening high-producing antibody-secreting cell from heterogeneity mammalian cell line populations is the vital procedure during the long time culture for antibody manufacture. To acquire high-producing cells, we designed a novel microfluidic chip for screening high-producing hybridomas from heterogeneous population at single cell level. All the procedures of hybridoma screening, including single cell trapping, identification, culturing and releasing, are integrated in a single chip. Compared with traditional methods based on limiting dilution, our chip realized higher monoclonal antibody (mAb) yield and shortened the screening period from 10 to 5 days with twice. It is significance that provides foundation to pharmaceutical company of rapidly screening cell lines.