A new Nd:YAG laser based single-photon fluorescence decay apparatus has been assembled and tested. The mode-locked Nd:YAG laser output is used to pump a dye laser whose doubled output is ideal for excitation of the ultraviolet fluorescence of proteins. Coupled with an automated sample changer and a new minicomputer, the instrument will represent the state-of-the-art in following fluorescence and anisotropy decay, as well as time-resolved emission spectra. The present system has been applied to problems in biochemistry involving the enzyme glutamine synthetase, the interaction of melittin and calmodulin, bovine serum albumin, liposome-dye complexes, and excited state protonation of serotonin. An atlas of fluorescence properties of about 30 protein dyes has been prepared and published. Various fluorescent or fluorogenic labels which react covalently with proteins have been attached to ovalbumin, and the excitation and emission spectra obtained. The spectra were then corrected for lamp and detector nonlinearities with wavelength. The atlas also contained fluorescence lifetime data on each dye-ovalbumin complex. The compendium of spectral and lifetime data the only one presently available of its kind.
