A Coumarin-Based Array for the Discrimination of Amyloids.

Self-assembly of misfolded proteins can lead to the formation of amyloids, which are implicated in the onset of many pathologies including Alzheimer's disease and Parkinson's disease. The facile detection and discrimination of different amyloids are crucial for early diagnosis of amyloid-related pathologies. Here, we report the development of a fluorescent coumarin-based two-sensor array that is able to correctly discriminate between four different amyloids implicated in amyloid-related pathologies with 100% classification. The array was also applied to mouse models of Alzheimer's disease and was able to discriminate between samples from mice corresponding to early (6 months) and advanced (12 months) stages of Alzheimer's disease. Finally, the flexibility of the array was assessed by expanding the analytes to include functional amyloids. The same two-sensor array was able to correctly discriminate between eight different disease-associated and functional amyloids with 100% classification.

Dual-Functionalisation of Fluorophores for the Preparation of Targeted and Selective Probes.

A key current challenge in biological research is the elucidation of the that roles chemicals and chemical reactions play in cellular function and dysfunction. Of the available cellular imaging techniques, fluorescence imaging offers a balance between sensitivity and resolution, enabling the cost-effective and rapid visualisation of model biological systems. Importantly, the use of responsive fluorescent probes in conjunction with ever-advancing microscopy and flow cytometry techniques enables the visualisation, with high spatiotemporal resolution, of both specific chemical species and chemical reactions in living cells. Ideal responsive fluorescent probes are those that contain a fluorophore tethered to both a sensing unit, to ensure selectivity of response, and a targeting group, to control the sub-cellular localisation of the probe. To date, probes that are both targeted and selective are relatively rare and most localised probes are discovered serendipitously rather than by design. A challenge in this field is therefore the identification of suitable fluorophore scaffolds that can be readily attached to both sensing and targeting groups. Here we review current strategies for dual-functionalisation of fluorophores, highlighting key examples of targeted, responsive probes.

Synthesis of Nitro-Aryl Functionalised 4-Amino-1,8-Naphthalimides and Their Evaluation as Fluorescent Hypoxia Sensors.

Fluorescent sensors are a vital research tool, enabling the study of intricate cellular processes in a sensitive manner. The design and synthesis of responsive and targeted probes is necessary to allow such processes to be interrogated in the cellular environment. This remains a challenge, and requires methods for functionalisation of fluorophores with multiple appendages for sensing and targeting groups. Methods to synthesise more structurally complex derivatives of fluorophores will expand their potential scope. Most known 4-amino-1,8-naphthalimides are only functionalised at imide and 4-positions, and structural modifications at additional positions will increase the breadth of their utility as responsive sensors. In this work, methods for the incorporation of a hypoxia sensing group to 4-amino-1,8-naphthalimide were evaluated. An intermediate was developed that allowed us to incorporate a sensing group, targeting group, and ICT donor to the naphthalimide core in a modular fashion. Synthetic strategies for attaching the hypoxia sensing group and how they affected the fluorescence of the naphthalimide were evaluated by photophysical characterisation and time-dependent density functional theory. An extracellular hypoxia probe was then rationally designed that could selectively image the hypoxic and necrotic region of tumour spheroids. Our results demonstrate the versatility of the naphthalimide scaffold and expand its utility. This approach to probe design will enable the flexible, efficient generation of selective, targeted fluorescent sensors for various biological purposes.

A fluorescent naphthalimide NADH mimic for continuous and reversible sensing of cellular redox state.

A fluorescent, naphthalimide-based, NADH mimic has been synthesised as a reversible, biocompatible, "on-off" probe for the detection of changes in intracellular redox environment (both oxidation and reduction). Interconversion was confirmed by means of electrochemistry and also 1H NMR, UV-vis and fluorescence spectroscopy. The reversibility was also successfully detected in A549 cells under simulated redox stress.