Oxygen molecules are essential for producing energy in living organisms, meaning that oxygen measurement is required not only in the medical field, but also in research experiments. Porphyrin dyes in polystyrene microparticles were loaded and embedded in dimethylpolysiloxane to create oxygen sensing film. The lifetime of phosphorescence emitted from the film by laser excitation varied depending on the oxygen concentration, and the oxygen partial pressure was quantified from the lifetime. We revealed that the smaller the microparticle size and film thickness, the shorter the response time to the change in oxygen concentration. Next, the film was placed in cell culture microfluidic devices that can form oxygen gradients in the culture layer, and changes of the gradients due to oxygen metabolism of cells and implanted tumor tissues on the sensing film. From this, we demonstrated the effectiveness of the developed oxygen sensing film for medical and biological applications.