Ink spreading and lateral light scattering in the substrate a↵ect the color of a halftone print. One of the most important phenomena which a↵ects the print result is dot gain, meaning that printed dots appear larger than the dots in the digital bitmap. This is partly due to the ink spreading and ink penetration into the substrate, resulting in an enhancement of the physical dot size, referred to as the physical dot gain. Lateral propagation of light in paper, causes printed dots to appear larger than their physical size, which is called optical dot gain. Characterization of total dot gain, i.e. the combination of physical and optical dot gain, is an important issue in the study of paper properties and print characteristics. Many models based on macroscopic measurements are reported in the literature to separately characterize both physical and optical dot gains. The aim of this study is to go beyond the macroscopic models, and to study the halftone prints on a microscopic scale, by using microscale images captured by a high-resolution camera.In this dissertation, three approaches based on the Murray-Davies model are proposed to obtain the total dot gain. In the first approach, by minimizing the root-mean-square di↵erence between the calculated spectrum and the reflected spectrum measured by the spectrophotometer, the total dot gain is approximated. The other two approaches are based on microscale images captured by a high-resolution camera. These two approaches di↵er in their schemes on how to obtain the gray tone of the full tone ink. By the use of microscale images, it is also possible to illustrate the shape of the e↵ective dot area for the investigated paper substrate.A novel approach based on the histogram of microscale images is also proposed to separate physical from optical dot gain. Attaining the physical dot gain characteristic makes it possible to determine the actual physical dot shape, by which the Modulation Transfer Function (MTF) of the paper substrate is estimated. The proposed approach is validated by comparing the estimated MTF of eleven o↵set printed coated papers to the MTF obtained from the unprinted papers using measured and Monte-Carlo simulated edge response.Another potential usage based on the separation of physical from optical dot gain, is to study the characterization of di↵erent color inks.ii