As-grown undoped zinc oxide (ZnO) films have been annealed in zinc-rich, oxygen-rich and vacuum ambient, and the electron concentration varied greatly after the annealing process. It decreased nearly two orders of magnitude after the sample was annealed in oxygen, while increased nearly three times after annealed in metallic zinc ambient, and increased slightly after annealed in vacuum. It was found that the variation trend of the electron concentration is always the same with the expected variation of oxygen vacancy (V ) under the three investigated conditions, it is thus speculated that V may be the dominant donor source in ZnO. By supplying more oxygen during the growth process to suppress V , ZnO films with lower electron concentration were obtained, which verifies the above speculation.