Antibiotic contamination and abuse are universal phenomena, accordingly antibiotic resistance has emerged as a serious issue globally in recent years. To detect antibiotics and their residues in the environment, foods, drugs and biological samples, numerous analytical techniques have been developed. Among them, aptamer-based methods are considered to be highly sensitive and selective and have been applied to the detection of various antibiotics in different samples. We present a systematical and critical review on the antibiotic-specific aptamers and their application in detection of antibiotics in different matrices, focusing on the recent advances in optical and electrochemical aptasensors, aptamer-affinity based sample purification, as well as the promising label-free and multiplex determination.