Cell behavior is regulated by transient interactions between biomolecules. Our understanding of cell machinery and of biological processes resides on the ability to detect such changes with high spatio-temporal resolution. Fluorescent reporters, able to effectively signal transient interactions between biomolecules with fluorescent microscopy techniques, provide a powerful means to detect these events with high efficiens.

An ideal fluorescent probe should be endowed with the following properties:

• Ease of synthesis and of functionalization;
• High sensitivity to the environment;
• High brightness and photophysical stability.

Our research is directed to the development of solvatochromic fluorescent probes able to report on polarity and viscosity changes in specific cell subdomains. Two classes of solvatochromic dyes are being investigated: switch-like and classical probes. Switch-like fluorophores can signal molecular recognition events at nanoscale by boosting their brigthness under certain environmental conditions (e.g. polarity changes during molecular recognition events). Classical solvatochromic dyes provide detailed ratiometric maps of polarity and dielectric constant at subcellular level. Furthermore, efficient probes for ratiometric fluorescence detection of viscosity and ordered/disordered lipid phases are being developed. Ongoing research involves engineering of our probes to obtain red-shifted absorption and fluorescence, as well as applicability to superresolution imaging techniques. Ideally, the engineered dyes will absorb and emit in the "biological optical window", thus allowing for more efficient signaling during in vivo studies.

Fig.1: Development of polarity fluorescent probes: from screening in cuvette (left, fluorescence in different solvents) to polarity measurements in living cells (center). The probe can be conjugated to biomolecules (e.g. lipids) and selectively vehiculated in specific domains