A microfluidic calorimeter was developed to measure dynamic temperature changes during glucose oxidation in the presence of layer-by-layer self-assembled glucose oxidase (GOD). The device contains a single flow channel which is 12xA0;mm wide, 60 mm long and 100 渭m high with two fluid inlets and one fluid outlet. An antimony-bismuth thin film thermopile with high common mode rejection ratio is attached to the lower channel wall of the microfluidic calorimeter. Layer-by-layer (LBL) assembly is used to form multilayers of glucose oxidase on the glass coverslips by alternate electrostatic adsorption of polyelectrolyte and enzyme. The heat released (鈭?9 kJ mol鈭?) by the oxidation of glucose in the presence of glucose oxidase is measured by the thermopile to detect the glucose concentration. Results showed an increased response with an increase in the number of immobilized enzyme layers or increase in the flow rate. The device has a lowest detection limit of 1 mg dL鈭? for glucose.