De novo design of dual-detection fluorescent sensors for bisulfite and copper (II) ion based on cascade activation

Cascade activated probes have received extensive attention due to their sequence-dependent detection of multiple markers in pathologic pathways. HSO₃^- and Cu²⁺ are biomarkers of intracellular mitochondrial activity, and their simultaneous detection is of great significance. Until now, probes that detect them simultaneously were all based on displacement mechanism with "ON-OFF-ON" switches in an emission channel. In other words, using HSO₃^- and Cu²⁺ as two inputs in logic sensing, the inputs of (0,0), (0,1), or (1,1) gave the same "1" output. Therefore, they suffered from the false positive signals in clinical application.

The de novo design of fluorescent probes with a more advanced sensing mechanism was explored. The four dyes, CM-Cu, CMA-Cu, R1, and R2, have similar structures, but their sensing behaviors are quite different. For dual detection of HSO₃^- and Cu²⁺, CM-Cu showed emission "ON-OFF-ON" switches based on IMP logic gate; while CMA-Cu, having one more aldehyde, was cascade activated based on AND gate for the first time. Noteworthily, unlike most dual-locked probes, which usually involve a complex synthesis process, here, cascade activation by HSO₃^- and Cu²⁺ is achieved by adjusting the electron distribution of a ready-made structure. Coupled with its low toxicity, high specificity, strong solid emission, and anti-diffusion ability, the sensing property of CMA-Cu have been validated in digital encryption and cell imaging.

We exhibited how to adjust the dual recognition from displacement to cascade activation by fine-tuning the molecular structure. This study may provide a new idea for the simple design of dual-locked probe, which is helpful for the sequential detection of multiple biomarkers with physiological correlation and could improve the accuracy of disease diagnosis.