Monoclinic Bi2O3:Ho3+ powder was synthesized using a co-precipitation method, followed by the deposition of Bi2O3:Ho3+ thin films on Si (100) substrates at various substrate temperatures (room temperature–600 °C) and oxygen partial pressures (5–200 mT) using pulsed-laser deposition. X-ray diffraction analysis showed a single α-Bi2O3 phase at temperatures of 400 and 500 °C, while a mixed α- and β-Bi2O3 phase was obtained at 600 °C. The films deposited at the different oxygen partial pressures showed an α-Bi2O3 and non-stoichiometric phase. The influences of different substrate temperatures and oxygen partial pressures on the morphology and the thickness of the films were analyzed using a scanning electron microscope. The root mean square roughnesses of the films were determined by using an atomic force microscope. The surface components, oxidation states and oxygen vacancies in all the deposited thin films were identified by X-ray photoelectron spectroscopy. The optical band gap of the Bi2O3:Ho3+ thin films was calculated using diffused reflectance spectra and was found to vary between 2.89 and 2.18 eV for the deposited films at the different temperatures, whereas the different oxygen partial pressures showed a band gap variation between 2.97 and 2.47 eV. Photoluminescence revealed that Ho3+ was the emitting centre in the isolated thin films with the 5F4/5S2 → 5I8 transition as the most intense emission in the green region.