The distribution and interactions of single molecules in three-dimensionally organized cellular networks are fundamental to the function of living systems. Today, we still lack a complete understanding of how local molecular mechanisms are integrated and dynamically mapped over larger scales onto functional activities. The challenges pose high demands for imaging technologies to provide molecular specificity, nanometer-scale resolution, ultrafast speed, and accessibility across larger volumes of tissues. In this presentation, I will talk about my laboratory’s recently developed super-resolution and light-field microscopy, and functional imaging tools for high-throughput extraction of molecular information in cells and tissues with ultrahigh-spatiotemporal resolution and accessibility.