An effective signal amplifying strategy for copper (II) sensing by using in situ fluorescent proteins as energy donor of FRET

Fluorescence resonance energy transfer (FRET) is a reliable, sensitive, and robust assay method for detection of many biological targets. However, it generally needs an externally-introduced label to form the donor-acceptor pair, which could alter the accuracy of the detection. To address this issue, we report herein the FRET-based reactive copper ion sensors by using in situ fluorescent proteins (FP) of human urine or blood serum as the energy donor. Using Bull Serum Albumin (BSA) as model proteins, the sensor exhibits a remarkably fluorescence enhancement when BSA binds to the surface of copper clusters (Cu NCs) base on electrostatic interaction. Conversely, low fluorescence enhancement is observed without using BSA. As proof-of-principle, this positive approach is directly applied to detect Cu²⁺ using urine as the energy donor, accompanying with a signal enhancement by two factors and low detection limit of 0.5 μM Cu²⁺. Moreover, the proposed sensor could be applied in other complex environments, such as blood serum or cell culture medium. Consequently, the effectiveness, simplicity and diversity of our proposed strategy enable the development of a class of probes toward complex human environment for rapid, sensitive, and selective detection of targets.