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Published byJair Moring Modified about 1 year ago

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More Non-linear Microscopy

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E x = E xo cos(2 ft) = E xo cos ( t) P x = aE xo cos ( t) + dE 2 xo cos 2 ( t) P = aE + dE 2 + d'E Nonlinear Polarization

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First experimental observation of SHG (1962) Right after invention of ruby laser

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Coherent Anti-Stokes Raman Scattering (CARS)

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Collect SHG, TPEF Simultaneously Laser scanning Nonlinear optical microscope 850 nm SHG is coherent: Forward propagates Or TPEF

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Evans and Xie Video Rate epi- CARS for Tissue Imaging

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Intensity ~ Power 2 Intensity ~ concentration Lifetime ~ ns Stoke shift Intensity ~ Power 2 Intensity ~ concentration 2 Instantaneous No loss of energy TPEF and SHG Photophysics

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Mertz, BJ )SHG “spectrum” mirrors Two-photon spectrum: Resonance enhancement 2) TPEF intensity >> SHG TPEF TPEF and SHG “Spectra”

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Only forward direction for objects axially longer than wavelength Dipole Emission Patterns for scattering particles Xie J. Phys. Chem Many dipoles aligned: All forward directed

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Mertz, BJ 2001 Calculations show that SHG emitted in two lobes

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WT OI/OI Fw Bw Fw Bw Forward/Backward Fractions SHG in Tendon: Wild type vs OI WT larger fibers, also brighter, F/B Ratio?

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image plane A Percent Scattered Back Thickness (µm) B Thick tissues, initial photon is multiply scatttered Some goes backwards direction Like SHG Backscattered collection Red, Black: experimental and Monte Carlo simulations From intralipid phantom Blue=mouse Real Tissue: Only Scattering Matters

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100 m 50 m 100 m 20 m 30 m deep 70 m deep 100 m deep 0 m deep ABC D E F 50 m CorneacytesSebaceous glands Adipocytes Evans and Xie PNAS CARS Image of Hairless Mouse Ear

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