We report a method to measure DNA hybridization potentiometrically in a manner conducive to portable or hand-held biosensors. An electrolyte-gated transistor (EGT) based on poly(3-hexylthiophene) (P3HT) and an ion-gel serves as a transducer for surface hybridization of DNA. The key aspect of the design is the use of a floating-gate electrode functionalized with ssDNA whose potential is determined by both capacitive coupling with a primary, addressable gate electrode and the presence of adsorbed molecules. When DNA is hybridized at the floating gate, it offsets the primary gate voltage felt by the P3HT semiconductor; the offset is directly measurable and quantitatively related to the number density of dsDNA molecules. The presented sensing strategy can be readily adapted to other biomolecules of interest and integrated into a microfluidic system for field applications of biosensors.