By virtue of its tunable electrical conductivity, remarkable solution processing capability, and great biocompatibility, polyaniline (PANI) has been recognized as an attractive active material for use in biological and chemical ("bio/chemical") sensors. This Spotlight article focuses on the structure and characteristics of PANI-based materials and the corresponding device-level sensing performance. PANI-based bio/chemical sensors, such as chemi-resistive sensors, electrochemical sensors, and transistor-based sensors, are systematically elucidated based on device architecture and sensing mechanism. The corresponding structure−function relationships among PANI doping state, microscopic structure, local crystallinity, and their functionalities in three types of sensors have been systematically elaborated. Finally, the state-of-the-art progress in applications of these sensors and the corresponding breakthroughs in a broader context are outlined from the challenges to strategies.