Organic thin‐film transistors (OTFTs) show promising applications in various chemical and biological sensors. The advantages of OTFT‐based sensors include high sensitivity, low cost, easy fabrication, flexibility and biocompatibility. In this paper, we review the chemical sensors and biosensors based on two types of OTFTs, including organic field‐effect transistors (OFETs) and organic electrochemical transistors (OECTs), mainly focusing on the papers published in the past 10 years. Various types of OTFT‐based sensors, including pH, ion, glucose, DNA, enzyme, antibody‐antigen, cell‐based sensors, dopamine sensor, etc., are classified and described in the paper in sequence. The sensing mechanisms and the detection limits of the devices are described in details. It is expected that OTFTs may have more important applications in chemical and biological sensing with the development of organic electronics.