1,721,562 research outputs found
Electric field depolarization in high aperture focusing with emphasis on annular apertures
No full textElectromagnetic focusing theory of light predicts that at high apertures field components arise that are polarized perpendicular to the initial polarization. Although vectorial depolarization has received considerable attention in focusing theory, no evidence has been presented as to its relevance in experiments. We measure the intensity of the perpendicularly orientated field in the focal region by utilizing monomolecular, fluorescent polydiacetylene layers whose transition dipoles are orientated in a single direction. For a 1.4 numerical aperture oil objective lens illuminated with linearly x-polarized light, we find that the integral of the modulus squared of the y-polarized focal field amounts to 1.5% of its x-polarized counterpart. In particular, we show here that the depolarization increases when using annular apertures. Annuli formed by a central obstruction with a diameter of 89% of that of the entrance pupil raise the integral to 5.5%. This compares well with the value of 5.8% predicted by electromagnetic focusing theory; however, the depolarization is also due to imperfections connected with focusing by refraction. Besides fluorescence microscopy and single molecule spectroscopy, the measured intensity of the depolarized component in the focal plane is relevant to all forms of light spectroscopy combining strong focusing with polarization analysis
4Pi-confocal images with axial superresolution
No full textWe present two-photon excitation 4Pi-confocal images of clustered fluorescence beads demonstrating three-dimensional far-field light microscopy with unprecedented resolution. For an excitation wavelength of 760 nm, the lateral and axial resolution amounts to 200 and 145 nm, respectively. The four-fold improved axial resolution is achieved by engineering the point-spread function through a suitable combination of aperture enlargement, two-photon excitation, confocalization and three-point deconvolution. In contrast to their confocal counterparts, 4Pi-confocal images do not exhibit the typical axial elongation, The axial resolution in the 4Pi-confocal images corresponds to about one-fifth of the wavelength and surpasses the lateral resolution by 25%
Depolarization by high aperture focusing
No full textWr introduce a method employing ferroelectric monomolecular layers, by which it is possible to measure the light field polarization in the focus of a lens. This method allowed us to noninvasively establish the perpendicularly oriented focal field that is anticipated at high apertures. For a numerical aperture 1.4 oil immersion lens illuminated with linearly polarized plane waves, the integral of the modulus square of the perpendicular component amounts to (1.51 +/- 0.2) % of that of the initial polarization. It is proven that depolarization decreases with decreasing aperture angle. Whereas for regular imaging conditions depolarization is largely negligible, it plays a significant role in microscopy of highest resolution, microspectroscopy, and single molecule studies
Increasing the Resolution of Far-Field Fluorescence Microscopy by Point-Spread-Function Engineering
No full textIncreasing the Resolution of Far-Field Fluorescence Microscopy by Point-Spread-Function Engineering
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Ground-state depletion fluorescence microscopy, a concept for breaking the diffraction resolution limit
No full textWe introduce and study a novel concept in farfield fluorescence microscopy fundamentally overcoming the classical diffraction resolution limit. This is accomlished by reducing the spatial extent of the effective focus of a scanning fluorescence microscope. The reduction is achieved by depleting the ground-state energy of the molecules located in the outer region of the focus. Our theoretical study shows that ground-state-depletion fluorescence microscopy has the potential of increasing the resolution of far-field fluorescence microscopy by an order of magnitude which is equivalent to a lateral resolution of 15 NM
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