Novel imaging technologies such as single-particle tracking provide tools to study the intricate process of virus infection in host cells. In this chapter, we provide an overview of studies in which single-particle tracking technologies were applied for the analysis of the viral entry pathways in the context of the live host cell. Single-particle tracking techniques have been dependent on advances in the fluorescent labeling microscopy method and image analysis. The mechanistic and kinetic insights offered by this technique will provide a better understanding of virus entry and may lead to a rational design of antiviral interventions.