Fluorescence emission detection using LED as excitation source

Abstract

We implement a Light Emitting Diode (LED)-excited fluorescence microscope coupled with a prism-grating-prism (PGP) dispersing element to quantitatively measure the emission wavelength of fluorescence spectra. We test our microscope on two common fluorescing samples, chlorophyll extracted from plant leaves and the topical antiseptic merthiolate. A fluorescence microscope with no dichroic beamsplitter is feasible for certain samples where both excitation and emission wavelengths are far from each other as in chlorophyll-b, which was excited at 470 nm and was observed to emit at 651.7 nm (0.26% error). Merthiolate which contains thimerosal, a mercury compound, was excited at 525 nm and gave a fluorescence spectrum with two peaks at 558 nm and 593 nm. The fluorescence emission was spectrally unmixed and was found to contribute 0.71 and 0.29 to the resultant spectrum, respectively. The shift from the 545 nm and 580 nm lines of elemental mercury is due to the fact that mercury exists as ions in the sample.