Selective Sensing of Catechol Aldehydes Levels in Living Systems using FLIM-FRET

Endogenous catechol aldehydes (CAs), namely 3,4-dihydroxyphenylacetaldehyde (DOPAL) and 3,4-dihydroxyphenylglycolaldehyde (DOPEGAL), play pivotal roles in neurobehavioral, cardiovascular, and metabolic processes. Dysregulation of CA levels contributes to neurological disorders and heart diseases. Thus, detecting imbalances in CAs levels is crucial for diagnosing early stages of CA-associated diseases. Here, we present innovative fluorescent sensors designed for rapid and selective detection of CAs within cells and tissue, overcoming the limitations of conventional diagnostic methods that necessitate cell destruction. The sensor operates by a dual-reaction trigger, leveraging the exceptional selectivity of o-phenylenediamine for aldehyde and phenylboronic acid for catechol, resulting in the production of a FRET signal exclusively for CAs in the presence of other aldehydes and catechols within cells. To circumvent issues such as spectral cross-talk, excitation intensity fluctuations, inner filtering, photobleaching, and detector sensitivity, we employed Fluorescence Lifetime Imaging Microscopy (FLIM) combined with FRET (FLIM-FRET) to accurately measure CAs levels at a nanosecond scale. This makes FLIM-FRET highly proficient for live cell and tissue imaging. Remarkably, we utilized this dual-reaction trigger FLIM-FRET system to detect endogenous CAs levels within cells in response to enzyme activators and inhibitors and within diseased-model mice tissue. These probes have the potential to serve as early warning systems for neurological diseases linked to CAs within living systems, laying the foundation for further investigations.