The FluoroDichroSpectroPhotometer: Multi-function Instrumentation for Biophysical Spectroscopy in the Ultraviolet, Visible and Near Infrare
AuthorsSutherland, John C.
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AbstractThe decade-wide region of the electromagnetic spectrum from wavelengths of roughly 150 nm in the far ultraviolet (UV) to 1.5 μ in the near infrared (NIR) is particularly important for biophysical spectroscopy because water is relatively transparent, while most other molecules important in bio-molecular systems absorb in some part of this "water window". In addition to spectrophotometers, which measure the absorption spectrum of a sample, the most widely used instruments are fluorometers, which measure the light emitted after absorption of a photon, and dichrometers, which measure the difference between the absorption or emission of two polarizations, either circular or linear. I will describe the rationales for, and the design and construction of a laboratory instrument capable of measuring fluorescence as well as circular dichroism (CD) and magnetic CD (MCD). A second instrument of the same general type was optimized for CD in the far and vacuum UV by the use of UV radiation from a synchrotron light source. Other developments have included the simultaneous measurement of CD and the absorption spectrum of the sample, fluorescence detected CD/MCD, and the measurement of fluorescence polarization anisotropy using the components normally associated with a dichrometer. Since its introduction in 1969, essentially all dichrometers have used photoelastic modulators (PEM) to periodically modulate the polarization of a monochromatic photon beam, which makes possible the detection of very small differences in the absorption of different polarization components of the same wavelength – to about one part in a million. I will also discuss the programming of PEMs as a function of wavelength to achieve their proper operation for the measurement of CD/MCD, linear dichroism (LD), and fluorescence polarization anisotropy, and the limits of certain approximations made in the derivation of the mathematical descriptions of the operations of dichrometers.
DescriptionPresentation given at the Materials Science Research Seminar Series on Friday, January 29, 2016, 1 – 2 PM.