A chemiluminescence assay for rapid and sensitive detection of Escherichia coli(E.coli)is developed,by using moving reaction boundary electrophoresis coupled with the luminol-H2O2-horseradish peroxidase system.The E.coli and its antibody is combined with horseradish peroxidase(HRP).Chemiluminescence was observed when HRP catalyze the reaction of the luminol-H2O2in the microchip channel,and a moving chemiluminescence reaction boundary(MCRB)was created during electrophoretic migration of the HRP.As the count of E.coli-antibody-HRP is in direct proportion to the velocity of MCRB,the E.coli contents can be read directly on a linear scale if the microchannel is graduated.More importantly,measurements of E.coliin replicate samples with MCRBmethod and a conventional ELISAs yielded the same quantitative results,including initial enzymatic reaction rates and enzyme activity.This methodeliminates the need of any fluorescence equipmentin chemiluminescence determination of E.coli for point-of-care diagnostics.The results demonstrate that MCRB in microchip electrophoresis is an easily accessible readout platform for quantitative and real-time bioanalyses.