Measurement of the Fluorescence Quantum Yield Using a Spectrometer With an Integrating Sphere Detector

Published

January 1, 2008

Author(s)

Adolfas K. Gaigalas, Lili Wang

Abstract

A method is proposed for measuring the fluorescence quantum yield (QY) using a commercial spectrophotometer with a 150 mm integrating sphere (IS) detector. The IS detector is equipped with an internal cuvette holder so that absorbance measurements can be performed with the cuvette inside the IS. In addition, the spectrophotometer has a cuvette holder outside the IS for performing conventional absorbance measurements. It is shown that the fluorescence quantum yield can be obtained from a combination of absorbance measurements of the buffer and the analyte solution inside and outside the IS detector. Due to the simultaneous detection of incident and fluorescent photons, the absorbance measurements need to be adjusted for the wavelength dependence of the photomultiplier detector and the wavelength dependence of the IS magnification factor. An estimate of the fluorescence emission spectrum is needed for proper application of the wavelength dependent adjustments. Results are presented for fluorescein, quinine sulfate, myoglobin, rhodamine B and erythrosin B. The QY of fluorescein was determined as 0.90 0.02 where the uncertainty is equal to the standard deviation of three independent measurements. The method provides a convenient and rapid estimate of the fluorescence quantum yield. Refinements of the measurement model and the characteristics of the IS can in principle yield a accurate value of the absolute fluorescence quantum yield.

Citation

Journal of Research (NIST JRES) -

Volume

113

Issue

1

NIST Pub Series

Journal of Research (NIST JRES)

Pub Type

NIST Pubs

Citation

Gaigalas, A. and Wang, L. (2008), Measurement of the Fluorescence Quantum Yield Using a Spectrometer With an Integrating Sphere Detector, Journal of Research (NIST JRES), National Institute of Standards and Technology, Gaithersburg, MD (Accessed April 25, 2024)