The influence of annealing temperature on the optical and electrical properties, nanostructure and surface morphology of ITO thin films prepared by ion-assisted electron beam evaporation on the glass substrates has been studied. The resistivity and transmittance spectra were measured by a four-point probe method and spectrophotometer, respectively. The nanostructure and surface morphology were examined by X-ray diffractometer and atomic force microscopy. The results show that the ITO thin films with a thickness of 200 nm is amorphous. The crystallite size and optical band gap of ITO thin films increased after annealing in vacuum at different temperatures from 200 to 350 oC. At 350 oC, high quality crystalline thin films with a crystallite size of about 30 nm were obtained. The average optical transmittance was 84% in the visible range (400-700 nm) and the resistivity of 1.34 × 10-4 W-cm was obtained at a temperature of 350 oC.