A new borate polymer PAA-ran-PAAPBA that can specifically adsorb glucose was introduced in the glucose measurement based on surface plasmon resonance, and the high-precision specific detection of glucose concentration was realized. Six layers and twelve layers of borate polymer were respectively bound onto the SPR sensors through the layer-by-layer self-assembly binding method, and the effect of different layers of borate polymer on the glucose surface plasmon resonance measurement was studied. The experiment was conducted in the concentration range of 1~10 mg·dL-1 (interval Δ=1 mg·dL-1), 10~100 mg·dL-1 (interval Δ=10 mg·dL-1), and 100~1 000 mg·dL-1(interval Δ=100 mg·dL-1), experiment data was fitted by quadric curve and the fitting degree of refractive index difference ΔRU and glucose concentration was obtained. Results showed that the 12-layer-polymer sensor was better than the 6-layer-polymer sensor in the first two smaller ranges, and the measuring result was not significantly affected by layers in the third range, indicating that for the small concentrations increasing polymer layer can dramatically improve the measurement.