The continuing hurdle of developing foodborne pathogen detection techniques is that compromises must be made among simplicity, portability, speed, sensitivity, and quantitation. Herein, we fabricated quantum dot nanobeads (QDNS) by a layer-by-layer assembly of quantum dots on the surface of polymer nanospheres. QDNS exhibited higher fluorescence intensity than the quantum dots at the same particle number. Based on the quantum dot nanobeads as the signal reporter, a quantitative lateral flow immunoassay was demonstrated for Salmonella typhimurium detection with improved sensitivity, specificity and accuracy. A visual detection limit of 5 × 103 CFU mL-1Salmonella typhimurium within 10 min has been proved and demonstrated. Additionally, higher concentrations of non-Salmonella typhimurium bacteria have negligible effects on the detection of Salmonella typhimurium. The results of 50 single blind tests by 10 testers suggested that the assay exhibited 100% accuracy. The results illustrate that the assay provides a balance among simplicity, speed, sensitivity and accuracy, and it can be a favorable alternative for Salmonella typhimurium screening in various samples.