Improving the sensitivity of the competitive lateral flow immunoassay (LFIA) is important, given the increasing demands for the monitoring of chemical contaminants in food. The choice of nanosized marker is an essential task for improving the LFIA sensitivity. In this study, a CdSe/ZnS quantum dot (QD)-based LFIA combined with a portable reader was developed for rapid and quantitative detection of an antibiotic lincomycin (LIN). The performance of the proposed fluorescence LFIA was compared to the conventional gold nanoparticle (AuNP)-based LFIA realized with the same immunoreagents. The visual cutoff values were 10 ng/mL for AuNP-based LFIA and 20 ng/mL for QD-based LFIA. Furthermore, the instrumental limits of detection have been shown to be comparable for both nanosized markers and amounted to 0.4 ng/mL for AuNPs and 0.2 ng/mL for QDs, respectively. According to the results obtained, both LFIAs may be used for rapid, cost-effective, on-site testing of antibiotics, in particular LIN. However, the QD-based LFIA exhibits lowest limit of detection with the least immunoreagent consumption, that makes it economically beneficial.