Reversible Ratiometric NADH Sensing Using Semiconducting Polymer Dots

Reduced nicotinamide adenine dinucleotide (NADH) is a key coenzyme in living cells due to its role as an electron carrier in redox reactions, and its concentration is an important indicator of cell metabolic state. Abnormal NADH levels are associated with age‐related metabolic diseases and neurodegenerative disorders, creating a demand for a simple, rapid analytical method for point‐of‐care NADH sensing. Here we develop a series of NADH‐sensitive semiconducting polymer dots (Pdots) as nanoprobes for NADH measurement, and test their performance in vitro and in vivo. NADH sensing is based on electron transfer from semiconducting polymer chains in the Pdot to NADH upon UV excitation, quenching Pdot fluorescence emission. In polyfluorene‐based Pdots, this mechanism resulted in an on‐off NADH sensor; in DPA‐CNPPV Pdots, UV excitation resulted in NADH‐sensitive emission at two wavelengths, enabling ratiometric detection. Ratiometric NADH detection using DPA‐CNPPV Pdots exhibits high sensitivity (3.1 μM limit of detection), excellent selectivity versus other analytes, reversibility, and a fast response (less than 5 s). We demonstrate applications of the ratiometric NADH‐sensing Pdots including smartphone‐based NADH imaging for point‐of‐care use. A simple, rapid analytical method for NADH sensing is achieved by electron transfer from a Pdot to NADH upon UV excitation. Ratiometric NADH detection using DPA‐CNPPV Pdots exhibits high sensitivity, excellent selectivity versus other analytes, and a fast and reversible response. Smartphone‐based in vivo ratiometric imaging of NADH is performed to demonstrate the feasibility of point‐of‐care use.