Physics

Infrared

Laser

STED microscopy

Optics

Photon upconversion

Microscopy

Femtosecond

Luminescence

Autofluorescence

Materials science

Penetration depth

Photobleaching

Quantum mechanics

Stimulated emission

Optoelectronics

Ytterbium

Excitation

Fluorescence

Nature Communications

Multiphoton fluorescence microscopy (MPM), using near infrared excitation light, provides increased penetration depth, decreased detection background, and reduced phototoxicity. Using stimulated emission depletion (STED) approach, MPM can bypass the diffraction limitation, but it requires both spatial alignment and temporal synchronization of high power (femtosecond) lasers, which is limited by the inefficiency of the probes. Here, we report that upconversion nanoparticles (UCNPs) can unlock a new mode of near-infrared emission saturation (NIRES) nanoscopy for deep tissue super-resolution imaging with excitation intensity several orders of magnitude lower than that required by conventional MPM dyes. Using a doughnut beam excitation from a 980 nm diode laser and detecting at 800 nm, we achieve a resolution of sub 50 nm, 1/20th of the excitation wavelength, in imaging of single UCNP through 93 μm thick liver tissue. This method offers a simple solution for deep tissue super resolution imaging and single molecule tracking.

Multi-photon near-infrared emission saturation nanoscopy using upconversion nanoparticles