Initiated by graphene, two-dimensional (2D) layered materials have attracted much attention owing to their novel layer-number-dependent physical and chemical properties. To fully utilize those properties, a fast and accurate determination of their layer number is the priority. Compared with conventional structural characterization tools, including atomic force microscopy, scanning electron microscopy, and transmission electron microscopy, the optical characterization methods such as optical contrast, Raman spectroscopy, photoluminescence, multiphoton imaging, and hyperspectral imaging have the distinctive advantages of a high-throughput and nondestructive examination. Here, taking the most studied 2D materials like graphene, MoS2, and black phosphorus as examples, we summarize the principles and applications of those optical characterization methods. The comparison of those methods may help us to select proper ones in a cost-effective way.