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Hex 4 from 5/18/2009 * * * * * * * Duke University, Fitzpatrick Institute for Photonics BIOS Lab, Department of Biomedical Engineering Two-wavelength unwrapping for transmission-geometry quantitative phase microscopy Matthew Rinehart, Natan T. Shaked, Adam Wax Phase Microscopy Phase microscopy measures relative optical path delays Samples that are optically thicker than λ cause 2π rollovers in phase maps Δ? m = ? Make equations look good with the animations. Transmission Phase Microscopy Modified Mach Zehnder interferometer with an off-axis reference arm N.T. Shaked, M.T. Rinehart, and A. Wax, Dual-interference-channel quantitative-phase microscopy of live cell dynamics, Opt. Lett.?34, 767-769 (2009)? Off-axis Quadrature Projections Wollaston prism splits images by polarization interfering 45° light with circularly polarized light creates a 90° phase delay Bayer mosaic filter splits 532nm light to G channel and 633nm light to R channel Color CCD channels Predicted crosstalk: ~ 5% per channel Sony? ICX205AK Acquisition phase mapping Acquire 2 simultaneous interferograms, 90° out of phase, subtract to remove DC I1 – I2 Acquisition phase mapping Fit to find fringe pattern frequency, ?c, and the phase, α Hilbert transform to remove complex conjugate Demodulate spatial image information Take angle to find wrapped phase map Make the formulas look good 2π rollover 2π rollover can be removed using unwrapping algorithms Sharp changes in phase cannot be accurately resolved Algorithms are complex computationally intensive Imaging at 2 wavelengths extends measurement range 633nm 532nm ? 3.33μm Find out the timescale that unwrapping algorithms follow with increasing image size. Subtract wrapped phase maps add 2π where 0 Amplifies phase noise Use beat wavelength phase map as guide to add correct multiple of 2π Two-wavelength unwrapping UV cure optical adhesive n = 1.53 Steep edges at structure boundaries 3-6μm predicted height from exposure