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Popularization of super-resolution imaging techniques has allowed cell biologists to probe cell structure and function in previously unattainable detail. These methodologies continue to evolve, with new improvements that allow tailoring the available techniques to a particular need and application. This collection showcases primary research articles, reviews and protocols and highlights these recent developments by exemplifying the new, interesting applications of super-resolution microscopy as well as related tool development. We hope that this compilation of works will inspire future research with the aim to resolve outstanding challenges and further expand the utility of super-resolution imaging across biological and medical disciplines.