A novel electrochemical sensor for nitrite was fabricated by modifying a glassy carbon electrode (GCE) with a polyethyleneimine and carbon quantum dots (PEI-CQDs) composite. The PEI-CQDs were synthesized by a simple one-pot method. PEI was used not only as a conductive polymer, but also as a nitrogen source to improve the electrocatalytic property and conductivity of CQDs. Moreover, the amino group of PEI can prevent CQDs from aggregating as well as selectively detect nitrite. The physicochemical properties, electrochemical properties, and nitrite sensing performance of these PEI-CQDs were characterized by various techniques. The effects of the pH, mass ratio of nitrogen to carbon sources, and applied potential were studied to find the optimal conditions for nitrite sensing. Under optimized conditions, amperometric and differential pulse voltammetric methods were employed to detect nitrite to obtain the detection range and detection limit, respectively. It was found that the proposed sensor based on PEI-CQDs/GCE exhibited comparable sensing performance to previously reported nitrite sensors with a wide linear range (0–1000 and 20–380 μM) and a low detection limit (2.87 and 1.16 μM). On the basis of the characterization data and the reaction kinetics of nitrite oxidation, a possible mechanism for this sensor is discussed. Moreover, the proposed sensor displayed high selectivity, good reproducibility, and stability under long-term storage and was successfully applied to the detection of nitrite in real samples.