Glucose selective oxidizing enzyme of glucose oxidase (GOD) was immobilized on the carboxyl group (–COOH) functionalized porous polyimide (PI) film (PI-GOD film). The –COOH functionalized film was fabricated by the modified breath figure method (BF) by casting the PI solution under humid conditions containing KOH. The amine group of GOD was covalently bonded to the PI-COOH film under catalytic conditions for the PI-GOD film. The PI-GOD film induced the color change of the bromothymol blue (BTB) indicator solution from green to yellow by adding glucose due to the oxidation of glucose by GOD to gluconic acid acting as an acidic source. Amine-terminated pH-sensitive polymethacrylic acid polymer (PMAA) was additionally immobilized to the PI-GOD film for the PI-GOD-PMAA film. The morphology of the pore surface of the PI-GOD-PMAA film was changed by the addition of glucose due to the coil-to-globule transition of the pH-sensitive polymer by the production of gluconic acid as a result of the reaction between glucose and GOD. The insulin adsorbed on the film was released by adding glucose due to the morphology change of the pH-sensitive polymer. The detection sensitivity of glucose was increased by the pore-selectively immobilized GOD acting as a micro-reactor.