A reference pH sensor, the preparation and application thereof. The reference pH sensor is an extended gate field effect transistor (EGFET) structure and comprises a metal oxide semiconductor field effect transistor (MOSFET) on a semiconductor substrate, a sensing unit comprising a substrate, a solid-state conductive sensing layer on the substrate, and a polypyrrole layer on the solid-state conductive sensing layer, and a metal wire connecting the MOSFET and the sensing unit.