Are Upconverting Ln(3+) Based Nanoparticles Any Good For Deep Tissue Imaging With Retention Of Optical Sectioning?

An effective strategy is presented to make spherical Ln(3+) doped NaYF4 nanoparticles that show upconversion, with the aim of deep-tissue optical imaging. Upconversion is the conversion of two or more low-energy photons into one of higher energy, e. g. 980 nm to 545 and 680 nm and 980 nm to 800 nm. In order to avoid the formation of nanoparticles with an aspect ratio, we developed a strategy in which subsequent shells were grown on spherical seed nanoparticles. The last shell is undoped in order to improve the optical properties. In addition, a simple intercalation strategy involving the oleate ligands on the surface has been developed to make the nanoparticles dispersible in aqueous solutions and physiological buffers. Two-photon upconversion laser scanning microscopy (TPULSM) and two-photon upconversion wide-field microscopy (TPUWFM) have been tested for their suitability in deep-tissue imaging with retention of lateral and depth resolution (also called optical sectioning). TPULSM can be used up to similar to 600 mu m deep, but takes inordinately long times to acquire, which is due to the fact that the absorption cross section of Yb3+ is low, the quantum yield of the upconversion process are << 1%, and the Ln(3+) excited states are up to several hundreds of mu s. Hence UCNPs in general are not very bright (i.e. large emitted photon flux). The TPUWFM seems more promising because acquisition times are only several minutes, with depth profiling up to 400 mu m. We show the first optical sectioning with this technique in the brain of a mouse, through a thin shaved skull.