Streptomycin, a broad-spectrum aminoglycoside antibiotic, is widely used in both human and veterinary medicine. Its overuse and environmental accumulation pose serious health risks, underscoring the need for sensitive and selective detection methods. In this study, we developed a molecularly imprinted fluorescence sensor (MIP-FS) by integrating nitrogen-doped carbon quantum dots (N-CQDs) with molecularly imprinted polymers (MIPs) for the selective detection of streptomycin. The N-CQDs were synthesized via a green microwave-assisted method using mangosteen peel as a sustainable and low-cost carbon source, representing an alternative biomass-based route for nanomaterial production. The resulting N-CQDs@MIPs displayed excellent fluorescence properties, high photostability, and strong selectivity for streptomycin through a fluorescence enhancement mechanism. Under optimized conditions, the sensor achieved a low detection limit of 3.2 nM (1.8 ppb) and a wide linear range from 0.01 to 8.00 μM. It also exhibited high selectivity against structurally similar antibiotics and common interfering ions. The method was successfully applied to pharmaceutical and environmental water samples, yielding results consistent with LC-MS/MS. These findings highlight the potential of the developed sensor as a rapid, sensitive, and eco-friendly tool for streptomycin detection in complex matrices.