Development of a Steady-State Fluorescence Spectroscopy System and a Time-Resolved Fluorescence Spectroscopy System

Abstract

This document describes thoroughly the construction of two spectroscopic systems: One system for obtaining fluorescence spectra and another system for obtaining the fluorescence lifetime. The first system consists of obtaining the fluorescence spectrum and is constituted by a Czerny-Turner type monochromator, a photomultiplier tube (PMT), a system for modulating the excitation signal and a system for treating and filtering the fluorescent signal constituted by an electromechanical chopper, a lock-in amplifier and a fast response PMT; plus a set of lenses and a samples holder. The fluorescein dissolved in ethanol was used as a calibration standard, and an Nd:YAG laser source was used as a source of excitation, which emits in 532 nm, of continuous wave and stable power in time adjustable by current. On the other hand, a fluorescence time-resolved system was developed. This system consists of obtaining the fluorescence lifetime of different samples under study from a Voltage curve Vs Time obtained from the system and showing the decay behavior of the phenomenon. This has a pulsed laser unit as a source of excitation, emitting at 532 nm, 80 mJ of energy and with a pulse width of 4 ns. For this system, two semiconductor photodiode detectors with fast response have been built; both with a response of temporary uptake above 1 ns, thus providing obtaining fluorescence lifetime up to this value. Fluorescein was also used in this system as a standard sample, but in this case dissolved in a buffer-like solution; accompanied also by a brief study of the change in the average lifespan of fluorescence in function of the pH of the solution. The results obtained show that two automated systems, reproducible, accurate and of very low cost compared to the current ones, have been successfully implemented for the study of steady-state and time-resolved fluorescence spectroscopy.