Super-resolution microscopy enabled by high-efficiency surface-migration emission depletion

Abstract Nonlinear depletion of fluorescence states by stimulated emission constitutes the basis of far-field stimulated emission depletion (STED) microscopy1,2. Despite significant efforts over the past decade, achieving resolution below the diffraction limit at low saturation intensities by STED remains a major technical challenge3,4. By taking advantage of the surface quenching effect in NaGdF4:Yb/Tm nanocrystals, we report here high-efficiency emission depletion through nanocrystal surface migration. Under a dual-beam, continuous-wave, laser manipulation (975-nm excitation and 730-nm de-excitation), we achieved an emission depletion efficiency of over 95% and a depletion saturation intensity of 18.3 kW cm-2. Emission depletion by surface migration through Gd sublattices enables super-resolution microscopic imaging with an average lateral resolution of 20 nm and a Fourier ring correlation (FRC) resolution of 24-32 nm (depletion beam intensity, 1.09 MW cm-2). Our approach circumvents the fundamental limitation of high-intensity STED microscopy, providing autofluorescence-free, re-excitation-background-free imaging with a depletion saturation intensity over three orders of magnitude lower than conventional fluorophores. We also demonstrated super-resolution imaging of actin filaments of fixed Hela cells labelled with 8-nm Tm3+-activated nanoparticles. Combined with the outstanding photostability of lanthanide luminescence, surface migration emission depletion could provide a powerful mechanism for low-power, super-resolution imaging or tracking of biological dynamics without photobleaching.