There is a significant demand for small, portable, and inexpensive analytical devices, which can be used in a wide range of sensing applications (e.g., food monitoring, detection of chemical, biological poisoning agents, environmental monitoring, medical diagnostics, military defense, etc.). Sensors based on organic semi-conducting polymers, which are suitable for large-area, low-cost, flexible, and eventually single-use throwaway electronics, provide a unique opportunity in that sense. We report on low-operating voltage organic field-effect transistor devices, which can be used as sensors in electrolytes and liquid media, using a biofunctionalized, biocompatible, regioregular poly(3-hexylthiophene) semiconducting layer. Measurements in electrolytes and complex media relevant for cell analysis have shown that the devices can be directly used as ion-sensitive transducers and are suitable for in vitro biosensing applications. With the demonstration of biocompatible semiconducting polymeric devices, we have overcome a substantial hurdle for the realization of low-cost and mass-produced sensors, opening new possibilities of biological sensing using organic devices.