Lysosome-Targeting Fluorescence Sensor for Sequential Detection and Imaging of Cu²⁺ and Homocysteine in Living Cells

A conjugated polymer-based fluorescence sensor, namely, PTNPy, was constructed on the basis of a polythiophene scaffold coupled with dimethylpyridylamine (DPA) groups in side chains for the consecutive detection and quantification of Cu²⁺ and Hcy in a perfect aqueous medium. A dramatic fluorescence quenching of PTNPy by the addition of Cu²⁺ was observed in Tris-HCl buffer solution (2 mM, pH 7.4), demonstrating a quick (<1 min) and highly selective response to Cu²⁺ with a low limit of detection of 6.79 nM. Subsequently, the Cu²⁺-quenched fluorescence of PTNPy can be completely recovered by homocysteine (Hcy), showing excellent selectivity to Hcy over other competitive species such as cysteine and glutathione. Thanks to the low cytotoxicity and lysosomal targeting ability of PTNPy, it was further applied as an optical sensor for the sequential imaging of Cu²⁺ and Hcy in HeLa cells. More importantly, Hcy concentration was linearly related to the fluorescence intensity of PTNPy in living cells, demonstrating huge potential for real-time monitoring the fluctuation of Hcy levels in living cells.