Fluorophores for confocal microscopy: Photophysics and photochemistry

Abstract

Fluorescence is probably the most important optical readout mode in biological confocal microscopy because it can be much more sensitive and specific than absorbance or reflectance, and because it works well with epi-illumination, which greatly simplifies scanner design. These advantages of fluorescence are critically dependent on suitable fluorophores that can be tagged onto biological macromolecules to show their location, or whose optical properties are sensitive to the local environment. Despite the pivotal importance of good fluorophores, little is known about how rationally to design good ones. Whereas the concept of confocal microscopy is only a few decades old and nearly all the optical, electronic, and computer components to support it have been developed or redesigned in the last few years, the most popular fluorophores were developed more than a century ago (in the case of fluoresceins or rhodamines) or several billion years ago [in the case of phycobiliproteins and green fluorescent proteins (GFPs)]. © 2006, 1995, 1989 Springer Science+Business Media, LLC. All rights reserved.