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Total Internal Reflection Fluorescence Microscopy
Interactive Tutorials
Polarized Light Evanescent Intensities
The light intensity at a TIRFM interface is a function of the illumination angle of incidence and the polarization of the incident light. This interactive tutorial explores how evanescent field intensities vary as a function of critical angle and the refractive index of the glass medium.
To operate the tutorial, use the Glass Refractive Index slider to change the refractive index value from a range of 1.45 to 1.78, corresponding to the differences between fused quartz and sapphire, respectively. As the slider is translated, the curves in the tutorial window display a change in both the critical angle and the relative intensities. Note how the evanescent field intensities drop as the refractive index and corresponding critical angle are decreased.
The p and s evanescent intensities are illustrated in the tutorial window as a function of incident angle for transmitted light in the lower refractive index medium when passed through an interface composed of a medium of variable refractive index (n(1) = 1.45-1.78) and water or an aqueous buffer solution (n(2) = 1.33). These calculations assume a condition of total internal reflection and require a critical angle of that is dependent upon the refractive index of the glass medium. Intensity, plotted on the ordinate, is expressed as the ratio of evanescent intensity at the interface (z equals zero) to the incident intensity for each polarization angle. It is interesting to note that the evanescent intensities for both polarization orientations exhibit a range between one and five times that of the plane wavefront incident intensity for angles within 15 degrees of the critical angle.
Contributing Authors
Daniel Axelrod - Department of Biophysics, University of Michigan, 930 North University Ave., Ann Arbor, Michigan 48109.
John C. Long and Michael W. Davidson - National High Magnetic Field Laboratory, 1800 East Paul Dirac Dr., The Florida State University, Tallahassee, Florida, 32310.
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