The paper is to build a novel glucose biosensor by the application of chitosan and the gold nanoparticles. This article explores the effect of glucose solution concentration, pH and temperature on the response current of the sensor. The experimental results show the sensor has good linear relationship when in the temperature of 35.0℃, pH= 7.00, glucose concentration in 0.018~25.7 mg/L, being I=4.1356m+91.787μA, R2=0.9979, and the detection limit is 0.012mg/L. The sensor has the advantages of simple preparation, high sensitivity and quick detection.