In this work, a green and low-cost voltammetric sensor, poly(L-Ornithine) film modified glassy carbon electrode (PLO/GCE) was fabricated for the first time to detect thiamethoxam. The PLO/GCE was prepared by one-step electrodeposition, which is environmentally friendly, convenient and highly controllable. The modified electrode was characterized by electrochemical method, IR and SEM. Moreover, the redox character of thiamethoxam at proposed sensor was studied systematically. The PLO/GCE showed high sensitivity to thiamethoxam detection. Utilizing the linear sweep voltammetry, a wider linear range (5.0 × 10−8 mol l−1 −9.0 × 10−6 mol l−1) and lower detection limit (1.7 × 10−8 mol l−1) (S/N = 3) were achieved. In addition, the proposed method was successfully applied to the determination of thiamethoxam in pesticide.