An enzymatic glucose sensor was fabricated and integrated into a heterodyne refractometer. A heterodyne light source was incident into the sensor which the circular birefringence refractive index of sample was changed as the chemical reaction between glucose oxidase and glucose progressed. Therefore, the phase difference before and after chemical reaction could be measured by a lock-in amplifier with phase-lock technology. The optical configuration and measurement procedures indicated that the alignment of the measurement system was straightforward and that it reduced the misalignment effect. The phase stability of the proposed system was superior to 0.04 deg within 1 min. The glucose sensor can be reused 100 times while retaining favorable performance. The results indicated that the resolutions of the proposed method were ∼1.5 and 1.29  mg  /  dl for the glucose solution and serum-based sample (SRM 965b), respectively. The calibration curve revealed high linearity within the dynamic range of 0 to 300  mg  /  dl. Based on these findings, the proposed method has a potential to measure the whole blood glucose concentration for daily monitoring.