Photoacoustic imaging (PAI) is an emerging imaging modality that shows great potential for preclinical research and clinical practice. As a hybrid technique, PAI uniquely combines the advantages of optical excitation and of acoustic detection. Optical excitation provides a rich contrast mechanism from either endogenous or exogenous chromophores, allowing PAI to perform biochemical, functional, and molecular imaging. Acoustic detection benefits from the low scattering of ultrasound in biological tissue, enabling PAI to generate high-resolution images in both the optical ballistic and diffusive regimes. Accordingly, this hybrid imaging modality features high sensitivity to optical absorption and wide scalability of spatial resolution with the desired imaging depth. Over the past two decades, the photoacoustic technique has led to a variety of exciting discoveries and applications from laboratory research to clinical patient care. In biological research, PAI has become an irreplaceable tool, providing functional optical contrast with high spatiotemporal resolution. Translational PAI also attracted growing interest in clinical applications including tumor margin examination, internal organ imaging, breast cancer screening, and sentinel lymph node mapping, among others.