Super-resolution fluorescence microscopy has revolutionized the ability to observe biological structures at resolutions beyond the diffraction limit of light. This technique allows the visualization of subcellular structures and dynamic processes in living cells at the nanometer scale, providing previously unobserved details of biological structures and processes. The diffraction limit of conventional fluorescence microscopy, approximately 200–300 nm in the lateral direction and 500–700 nm in the axial direction, restricts the resolution of light microscopy. However, recent developments in super-resolution techniques have overcome this limitation, enabling three-dimensional imaging, multicolor colocalization, and dynamic processes in living cells. Several super-resolution techniques have been developed, including STED, RESOLFT, SSIM, STORM, PALM, and FPALM. These methods achieve a significant improvement in spatial resolution by utilizing nonlinear effects, structured illumination, or single-molecule localization. STED uses a second laser to suppress fluorescence emission from fluorophores outside the center of the excitation, resulting in a sharper PSF. RESOLFT employs photoswitchable fluorophores to achieve super-resolution by depleting fluorescence at the periphery of the excitation. SSIM uses structured illumination to enhance resolution by mixing spatial frequencies of the illumination pattern with those of the sample. STORM, PALM, and FPALM achieve super-resolution by localizing individual fluorescent molecules with high precision. These techniques have enabled the observation of previously unresolved details of cellular structures, demonstrating their potential in elucidating biological processes at the cellular and molecular scale. However, challenges remain in achieving high-resolution imaging in all three dimensions, multicolor imaging, and live-cell imaging. The resolution of these techniques is limited by factors such as the saturation level of fluorescence, the number of photons detected, and the labeling density. Additionally, the time resolution of super-resolution microscopy is generally slower than conventional fluorescence microscopy due to the need for high-precision localization of individual fluorophores. Despite these challenges, super-resolution fluorescence microscopy is becoming a widely used tool for cell and tissue imaging, providing unprecedented insights into biological structures and processes.Super-resolution fluorescence microscopy has revolutionized the ability to observe biological structures at resolutions beyond the diffraction limit of light. This technique allows the visualization of subcellular structures and dynamic processes in living cells at the nanometer scale, providing previously unobserved details of biological structures and processes. The diffraction limit of conventional fluorescence microscopy, approximately 200–300 nm in the lateral direction and 500–700 nm in the axial direction, restricts the resolution of light microscopy. However, recent developments in super-resolution techniques have overcome this limitation, enabling three-dimensional imaging, multicolor colocalization, and dynamic processes in living cells. Several super-resolution techniques have been developed, including STED, RESOLFT, SSIM, STORM, PALM, and FPALM. These methods achieve a significant improvement in spatial resolution by utilizing nonlinear effects, structured illumination, or single-molecule localization. STED uses a second laser to suppress fluorescence emission from fluorophores outside the center of the excitation, resulting in a sharper PSF. RESOLFT employs photoswitchable fluorophores to achieve super-resolution by depleting fluorescence at the periphery of the excitation. SSIM uses structured illumination to enhance resolution by mixing spatial frequencies of the illumination pattern with those of the sample. STORM, PALM, and FPALM achieve super-resolution by localizing individual fluorescent molecules with high precision. These techniques have enabled the observation of previously unresolved details of cellular structures, demonstrating their potential in elucidating biological processes at the cellular and molecular scale. However, challenges remain in achieving high-resolution imaging in all three dimensions, multicolor imaging, and live-cell imaging. The resolution of these techniques is limited by factors such as the saturation level of fluorescence, the number of photons detected, and the labeling density. Additionally, the time resolution of super-resolution microscopy is generally slower than conventional fluorescence microscopy due to the need for high-precision localization of individual fluorophores. Despite these challenges, super-resolution fluorescence microscopy is becoming a widely used tool for cell and tissue imaging, providing unprecedented insights into biological structures and processes.