AbstractAntibiotic abuse is considered as a serious problem affecting human health, demanding great attention to explore robust, accurate, real-time, on-site, and sensitive methods for rapid evaluation (detection and quantification) of food and biological samples such as serum. To address this challenging problem, biosensors have been developed as a valuable and sensitive tool to detect and quantify the amount of antibiotics. Among various kinds of biosensors, recently, aptamer-based biosensors (aptasensors) based on the fluorescent strategy have evolved as an excellent candidate for rapid evaluation of antibiotics, owing to their superior selectivity, specificity, and sensitivity. This review encompasses an overview of various kinds of recently developed fluorescent aptasensors for antibiotic detection and gives an idea of important sensing mechanisms associated with the developed aptasensors. The current review focuses on aims to further encourage and inspire targeted readers to develop new approaches for fabricating more practical and mature fluorescent aptasensors for antibiotic detection in the future.