Anovel chemiluminescence (CL) flowsensor for in vivo on-line monitoring of glucose in blood by microdialysis sampling is described in this paper. The sol-gel method is introduced to co-immobilize horseradish peroxidase (HRP) and glucose oxidase (GOD) on the inside surface of the CL flow cell. The CL detection involved enzymatic oxidation of glucose to d-gluconic acid and H2O2, then H2O2 oxidizing luminol to produce CL in presence of HRP. The perfusion rate of the dialysate was 5.0ml/min. The dialysate sample volume was 20ml, and the sample throughput was 15h−1. The variation of glucose level in blood of the rabbit was monitored after the administration of glucose to demonstrate the favorable resolution and reliability of the system for in vivo on-line monitoring.