by Anubhav Dhar, Sucheta Dey, Sanjana Mullick, Nishant Kumar Suman, Maxime C. van Zwam, Nishaant Kumar Palani Balaji, Angana Ghosh, Deepak Nair, Koen van den Dries, Sudarshan Gadadhar, Saravanan Palani Expansion microscopy (ExM) has revolutionized super-resolution imaging in cell biology due to its simple and inexpensive workflow. The use of ExM has revealed several novel insights into the nanoscale architectures of cellular protein complexes, especially the microtubule cytoskeleton in model and non-model systems. Despite tremendous progress in expansion microscopy protocols that preserve cellular ultrastructure (U-ExM), compatible probes for imaging actin isoforms with U-ExM are still lacking and have hindered the study of diverse actin isoforms and networks across model systems. Here, we use IntAct, an internally tagged actin that incorporates into cellular actin networks, to develop and optimize U-ExM for diverse actin structures in yeast, mammalian cells, and primary neurons. Using