Investigating the physics in tumor development and progression has been recognized as a promising way to advance both basic cancer research and clinical cancer treatment. The advent of atomic force microscopy (AFM) provides an exciting multifunctional tool for investigating the physical properties of biological systems under aqueous conditions with unprecedented spatiotemporal resolution, opening new possibilities for nanomedicine in physical oncology. In this article, the biomedical applications of AFM to investigate the physical sciences in oncology at single-cell, single-molecule, and tissue levels are systematically reviewed, taking lymphomas as an example. First, detecting the physics of cancers at multiple scales is summarized, along with antibody-based therapy for lymphomas. Next, the principles and methods of AFM imaging and measurements are summarized. After that, utilizing AFM to probe the physics of tumors is reviewed from several facets, including topographical imaging, mechanical measurement, molecular recognition, primary cell detection, and tissue characterization. In particular, the motivations and state-of-the-art overview of each facet is presented. Finally the current limitations and future directions of AFM assays for cancer investigations are discussed..