ISS Spectrofluorometry

ISS Spectrofluorometry –Steady State and Lifetime [Funded by NIH – PPG project with Dr. Ben Gaston, Pediatrics, SOM; 2017]

The Chronos is a compact and portable spectrofluorometer for fluorescence lifetime measurements with picosecond resolution and for the acquisition of steady-state fluorescence measurements (polarization measurements at fixed wavelengths, kinetics studies; xenon arc lamp 230-850nm). The basic sample compartment instrument is designed for lifetime fluorescence measurements with time-correlated single-photon counting (TCSPC) using the data acquisition by Becker and Hickl (BH); the instrument can be equipped for frequency-domain data acquisition as well. The instrument uses pulsed lasers (laser diodes, Ti:Sapphire laser) as light sources (Pulsed LEDs - 280 nm & 370 nm). Photomultiplier tube (Model Hamamatsu H7422P-40), GaAsP photocathode, QE=40% at the peak, wavelength range 300-720 nm.

The Vinci Multidimensional Fluorescence Spectroscopy Analysis software for Windows 7 controls the instrument automation and its interface to computer-controlled external devices (temperature bath, titrator). Vinci includes routines for the automatic acquisition of unidimensional data files (excitation and emission spectra; polarization and anisotropy spectra; synchronous luminescence spectra; slow and fast kinetics studies; lifetime data; rotational correlation times) and multidimensional data files (intensity and polarization versus excitation and emission wavelengths, time, temperature, lifetime). Raw data are stored in ASCII format along with the experimental parameters. The analysis portion of the software includes data manipulation (operations between spectra, smoothing, correction, derivative and integration). The analysis includes fitting routines for the determination of multiple decay times (exponential, non-exponential, lifetime distributions); for the determination of rotational correlation times (isotropic, anisotropic and hindered rotators); for the display of time-resolved spectra and the resolution of spectra using phase and modulation.