Photoactivatable Fluorophores

Françisco M Raymo

doi:10.5402/2012/619251

Abstract

Photoactivatable fluorophores switch from a nonemissive to an emissive state upon illumination at an activating wavelength and then emit after irradiation at an exciting wavelength. The interplay of such activation and excitation events can be exploited to switch fluorescence on in a defined region of space at a given interval of time. In turn, the spatiotemporal control of fluorescence translates into the opportunity to implement imaging and spectroscopic schemes that are not possible with conventional fluorophores. Specifically, photoactivatable fluorophores permit the monitoring of dynamic processes in real time as well as the reconstruction of images with subdiffraction resolution. These promising applications can have a significant impact on the characterization of the structures and functions of biomolecular systems. As a result, strategies to implement mechanisms for fluorescence photoactivation with synthetic fluorophores are particularly valuable. In fact, a number of versatile operating principles have already been identified to activate the fluorescence of numerous members of the main families of synthetic dyes. These methods are based on either the irreversible cleavage of covalent bonds or the reversible opening and closing of rings. This paper overviews the fundamental mechanisms that govern the behavior of these photoresponsive systems, illustrates structural designs for fluorescence photoactivation, and provides representative examples of photoactivatable fluorophores in actions.

Highlights

Certain organic molecules emit light in the form of fluorescence upon excitation at an appropriate wavelength [1]
The unique combination of photochemical and photophysical properties of photoactivatable fluorophores translates into the opportunity to control the spatial and temporal distribution of fluorescence
Synthetic fluorophores can be designed to emit light in the form of fluorescence upon excitation at one wavelength only after activation at another wavelength. Such photoactivatable fluorophores can be constructed by connecting a photocleavable group to a fluorescent chromophore

Summary

Introduction

Certain organic molecules emit light in the form of fluorescence upon excitation at an appropriate wavelength [1] This behavior offers the opportunity to probe biological structures and processes with optical methods [2]. Mechanisms to switch molecules from a nonemissive to an emissive state under the influence of optical stimulations translate into the opportunity to photoactivate fluorescence. Interest in such photoactivatable fluorophores was sparked initially by the idea of monitoring the course of photochemical reactions with fluorescence measurements [4] and by the possibility of using such compounds for photographic applications [5]. That photoactivatable fluorophores can be a valuable complement to conventional ones

Objective lens

Photophysical Properties of Organic Molecules

Mechanisms and Structural Designs for Reversible Fluorescence Photoactivation

Applications of Photoactivatable Fluorophores

Conclusions