New 4-styrylcoumarin Derivatives as Potentials Fluorescent Labels for Biomolecules: Application in RNA-fish Probes

Fluorescence microscopy is a highly sensitive imaging technique used in various scientific fields such as cellular biology, environmental sciences, medicine, and pharmacy. It offers the advantage of using multiple fluorescent labels to visualize different biomolecules and generate multicolored images for identifying specific components within complex biomolecular structures and studying their interactions. These fluorescent labels create chemically stable and minimally disruptive bioconjugates. Amino-reactive fluorescent labels, due to their ease of incorporation into biomolecules, are commonly used in applications like fluorescence in situ hybridization, histochemistry, cell tracing, receptor binding, and immunochemistry. However, the existing popular fluorescent labels are expensive, making coumarin derivatives a potential cost-effective solution for developing bright fluorophores. In this study, the fluorescent 4-styrylcoumarin derivative labels were synthesized and evaluated as potentially effective fluorescent labels for biomolecules. Twelve new fluorescent oligonucleotide probes have been obtained, 6 directed to the rRNA region of eukaryotic cells (EUK516) and 6 directed to the rRNA region of prokaryotic cells (EUB338). The developed fluorescent probes were tested on microorganisms belonging to the culture collection of the Laboratory of Biodegradation and Biotechnology of the HERCULES Laboratory (University of Évora, Portugal), showing effective performance as RNA-FISH probes. These findings evidenced the applicability of the new 4-styrylcoumarin derivatives in labeling of biomolecules and bioimaging.