Epifluorescence microscopy with a pulsed nitrogen tunable dye laser source.

Abstract

This paper describes an epifluorescence system that uses a pulsed laser source as an excitation source for quantitative fluorescence measurements. A 1 micron diameter portion of the sample can be illuminated with high intensity, short light pulses and the fluorescence can be measured with a fast detection system. Selective excitation is possible at 337 nm and from 357 to 710 nm by using different dyes in the laser system. The spectral bandwidth is 0.1-3 nm. Several advantages over the commonly used systems with continuum spectral sources are indicated, especially when measured intensities are low, as in the case of intrinsic fluorescence of the sample, and where reflection of unwanted excitation light may cause appreciable errors when using conventional light sources.