Recent developments on the preparation and application of ZnO films for acoustic wave-based microfluidics and biosensors are reviewed in this paper. High quality and strongly textured ZnO thin films can be prepared using many technologies, among which RF magnetron sputtering is most commonly used. This paper reviews the deposition of ZnO film and summarizes the factors influencing the microstructure, texture and piezoelectric properties of deposited ZnO films. ZnO acoustic wave devices can be successfully used as biosensors, based on the biomolecule recognition using highly sensitive shear horizontal and Love-wave surface acoustic waves, as well as film bulk acoustic resonator devices. The acoustic wave generated on the ZnO acoustic devices can induce significant acoustic streaming, small scale fluid mixing, pumping, ejection and atomization, depending on the wave mode, amplitude and surface condition. The potential to fabricate an integrated lab-on-a-chip diagnostic system based on these ZnO acoustic wave technologies is also discussed.