The setup
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• Our in-line holographic video microscope shown is based on an inverted microscope outfitted with a 100x numerical aperture 1.4 oil immersion objective.
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• The conventional incandescent illumination is replaced with the collimated coherent beam from either a He-Ne laser or a solid-state laser .
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• Individual colloidal spheres scatter a small proportion of the incident beam, and the scattered light interferes with the unscattered portion in the focal plane of the microscope's objective lens.
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• The microscope magnifies the interference pattern and projects it onto the face of a low-noise gray-scale video camera (NEC TI 324 IIA), with a total system magnification of 101 nm/pixel.
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• The video stream is recorded as uncompressed digital video with a digital video recorder (Pioneer H520S) for subsequent analysis.
The Summary of the analysis method
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3. Estimate the starting point of the fit (spheretool.pro)
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4. Loop the tracking procedure for all frames
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5. Link the coordinates found in each frame together to form trajectories.
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6. Analyze the trajectory info as you see fit for your particular experiment.