Macrophages play crucial roles in immune responses and tissue homeostasis. Despite the fact that macrophages were described more than a century ago, they continue to be the cells of intensive interest. Advanced understanding of phenotypic diversity in macrophages holds great promise for development of cell-based therapeutic strategies. The introduction of innovative approaches in cell biology greatly enhances our ability to investigate the unique characteristics of macrophages. The review considers both classical methods to study macrophages and high-tech approaches, including single-cell sequencing, single-cell mass spectrometry, droplet microfluidics, scanning probe microscopy and atomic force spectroscopy. This review will be valuable both to specialists beginning their study of macrophages and to experienced scientists seeking to deepen their understanding of methods at the intersection of biological and physical sciences.