BACKGROUND The recently developed super-resolution microscopy has been applied to characterize the nanometer-scale structure of actin filaments in live cells, owing to its live-cell imaging ability at sub-diffraction resolution. However, there are still some limitations to existing live-cell labeling methods for super-resolution imaging of F-actin. RESULTS A series of cell-permeable fluorescent probes was developed for super-resolution imaging of actin filaments with the introduction of a cell-penetrating peptide, (rR)(3)R-2. These probes are highly specific, multicolor available and suitable for different cell lines. With these probes, we resolve actin filaments at spatial resolution of ca 60 nm in live-cell super-resolution imaging. CONCLUSIONS Our results here extensively expand the types of probes for live-cell super-resolution imaging of F-actin and open up a new avenue in the design of cell-permeable fluorescent probes. (c) 2019 Society of Chemical Industry