Fig. 2.

The principle of NIRES nanoscopy using UCNP as multiphoton probe for deep tissue imaging. a The simplified energy levels and upconversion process of Yb³⁺ and Tm³⁺ co-doped UCNPs. The sensitizer Yb³⁺ ions initiate the photon upconversion process by a linear and sequential absorption of 980 nm excitation. Due to the multiple long-lived intermediate states, the energy is stepwise transferred onto the scaffold energy levels of emitters Tm³⁺, eventually facilitate multiphoton upconversion emission, including emissions from the four photon upconversion excited state ¹D₂ (455 nm), three photon state ¹G₄ (470 nm), and two photon excited state ³H₄ (800 nm). b The saturation intensity curve of the 800 nm emissions from UCNPs (40 nm NaYF₄: 20% Yb³⁺, 4% Tm³⁺) under 980 nm excitation. c Cross-section profiles of the saturated upconversion emission of UCNPs at four different excitation powers of 0.1, 0.4, 1, and 3 MW cm⁻². d The simulated “negative” contrast images of the cross-section profiles of a single UCNP. Pixel size, 10 nm. Scale bar is 500 nm