Unconjugated
A near-infrared fluorescent probe (DDAB) for highly selective and sensitive detection of carboxylesterase 2 (CE2) has been designed, synthesized, and systematically studied both in vitro and in vivo. Upon addition of CE2, the ester bond of DDAB could be rapidly cleaved and then release a near-infrared (NIR) fluorophore DDAO, which brings a remarkable yellow-to-blue color change and strong NIR fluorescence emission in physiological solutions. The newly developed probe exhibits excellent properties including good specificity, ultrahigh sensitivity and high imaging resolution. Moreover, DDAB has been applied to measure the real activities of CE2 in complex biological samples, as well as to screen CE2 inhibitors by using tissue preparations as the enzymes sources. The probe has also been successfully used to detect endogenous CE2 in living cells and in vivo for the first time, and the results demonstrate that such detection is highly reliable. All these prominent features of DDAB make it holds great promise for further investigation on CE2-associated biological process and for exploring the physiological functions of CE2 in living systems.
In this study, a two-photon ratiometric fluorescent probe NCEN has been designed and developed for highly selective and sensitive sensing of human carboxylesterase 2 (hCE2) based on the catalytic properties and substrate preference of hCE2. Upon addition of hCE2, the probe could be readily hydrolyzed to release 4-amino-1,8-naphthalimide (NAH), which brings remarkable red-shift in fluorescence (90 nm) spectrum. The newly developed probe exhibits good specificity, ultrahigh sensitivity, and has been successfully applied to determine the real activities of hCE2 in complex biological samples such as cell and tissue preparations. NCEN has also been used for two-photon imaging of intracellular hCE2 in living cells as well as in deep-tissues for the first time, and the results showed that the probe exhibited high ratiometric imaging resolution and deep-tissue imaging depth. All these findings suggested that this probe holds great promise for applications in bioimaging of endogenous hCE2 in living cells and in exploring the biological functions of hCE2 in complex biological systems.