Fluorescence Spectroscopy: A Powerful Tool for Biological Systems

Abstract

Fluorescence spectroscopy is one of the most widely used spectroscopic techniques in the fields of biochemistry and molecular biophysics today. Although Fluorescence measurements do not provide detailed structural information, the technique has become quiet popular because of its acute sensitivity to changes in the structural and dynamic properties of the biomolecules and biomolecular complexes. Like most biophysical techniques, fluorescence spectroscopic studies can be carried out at many levels ranging from simple measurement of steady state emission intensity to quiet sophisticated time-resolved studies. The four basic rules of fluorescence are as follows 1.The Frank- Condon principle: the nuclei are stationary during electronic transitions, and so excitation occurs to vibrationally excited levels of the excited electronic state. 2.Emission occurs from the lowest vibrational level of the lowest excited singlet state because relaxation from the excited vibrational levels is much faster than emission. 3. The stokes shift: emission is always of lower energy than absorption due to nuclear relaxation in the excited state. 4.The mirror image rule: emission spectra are mirror images of the lowest energy absorption band.