A simple and inexpensive method to fabricate a colloidal CdSe/ZnS quantum dots-modified paper-based assay for glucose is herein reported. The circular paper sheets were uniformly loaded and displayed strong fluorescence under a conventional hand-held UV lamp (365 nm). The assay is based on the use of glucose oxidase enzyme (GOx), which impregnated the paper sheets, producing H2O2 upon the reaction with the glucose contained in the samples. After 20 min of exposure, the fluorescence intensity changed due to the quenching caused by H2O2. To obtain a reading, the paper sheets were photographed under 365 nm excitation using a digital camera. Several parameters, including the amount of QD, sample pH, and amount of GOx were optimized to maximize the response to glucose. The paper-based assay showed a sigmoidal-shaped response with respect to the glucose concentration in the 5-200 mg∙dL-1 range (limit of detection of 5 µg∙dL-1), demonstrating their potential use for biomedical applications.