An N-nitrosation reactivity-based two-photon fluorescent probe for the specific detection of NO was rationally designed, prepared, and applied in the in situ detection of nitric oxide in ischemia reperfusion injury mouse model under two-photon microscopy for the first time.

Two-photon (TP) fluorescent probes are potential candidates for near-infrared (NIR) imaging which holds great promise in biological research. However, currently, most TP probes emit at wavelength <600 nm, which impedes their practical applications. In this work, we explored the TP properties of a silicon-rhodamine (SiR) derivative and hence developed the first SiR scaffold based "NIR-to-NIR" TP probe (SiRNO) for nitric oxide (NO). SiRNO exhibited high sensitivity and specificity, as well as fast response for NO detection. It was able to track the subtle variation of intracellular NO content in live cells. Owing to the NIR excitation and emission, SiRNO enabled the detection of NO in situ in the xenograft tumor mouse model, revealing the NO generation during the tumor progression. This work indicates that SiR can be an ideal platform for the development of NIR emissive TP probe and may thus promote the advancement of NIR imaging.

Peroxynitrite (ONOO À ) plays a critical role in Alzheimer's disease (AD). To reveal the ONOO À influx in AD brains, an activatable activity-based fluorescence probe Rd-DPA3 was designed by a structure-modulated strategy. Taking advantage of ONOO À -initiated twostep cascade reactions of a novel chemical trigger, Rd-DPA3 specifically responds to ONOO À in 0.3 mM of other reactive oxygen species (ROS) and varied proteins, and gives an intensive fluorescence enhancement (F/F 0 = 50). Moreover, with its mitochondria-targeting ability, Rd-DPA3 can be used to efficiently monitor the alternations of intracellular ONOO À levels in cerebral cells during oxidative stress. Significantly, due to NIR emission and good blood-brain barrier (BBB) crossing ability, Rd-DPA3 is suitable for in vivo imaging of cerebral ONOO À influx and illustrating an age-dependent accumulation of ONOO À in AD mice brains.

A ratiometric two-photon probe was designed to visualize HClO levels in tissues, revealing the generation of HClO in the wound-repairing process of mice for the first time.