Optical contrast is the most common preliminary method to identify layer number of two-dimensional (2D) materials, but it is seldom used as a confirmatory technique. We explain the reason for variation of optical contrast between imaging systems, motivating system-independent measurement of optical contrast as a critical need. We describe a universal method to quantify the layer number using the RGB (red–green–blue) and RAW optical images. For RGB images, the slope of 2D flake (MoS₂, WSe₂, graphene) intensity vs substrate intensity is extracted from optical images with varying lamp power. The intensity slope identifies layer number and is system independent. For RAW images, intensity slopes and intensity ratios are completely system and intensity independent. Intensity slope (for RGB) and intensity ratio (for RAW) are thus universal parameters for identifying layer number. The RAW format is not present in all imaging systems, but it can confirm layer number using a single optical image, making it a rapid and system-independent universal method. A Fresnel-reflectance-based optical model provides an excellent match with experiments. Furthermore, we have created a MATLAB-based graphical user interface that can identify layer number rapidly. This technique is expected to accelerate the preparation of heterostructures and to fulfill a prolonged need for universal optical contrast method.

Paper available at: https://doi.org/10.1021/acsnano.2c04833