Realization and Implementation of Two-photon Acoustic Microscopy
Date Issued
2015
Date
2015
Author(s)
Lee, Szu-Yu
Abstract
As a new branch of optical-absorption based microscopy with detection of non-radiative emission, photoacoustic microscopy provides manifold bio-information with scalable field of view inside biological specimens. For cell or organelle micro-level imaging, high resolving capability is necessary. However, among common photoacoustic microscopy, high lateral and axial resolution is achieved separately by different engineering methods. Furthermore, imaging rate is relatively slow compared to other in-vivo imaging modalities. To simultaneously obtain high spatial resolution in three dimensions, in this thesis, I report an in-vivo photoacoustic microscopy by a distinctive physical mechanism of contrast − two-photon absorption. The light source is a Ti:sapphire laser with wavelength at 800~860 nm. Due to the nonlinear absorption property, intrinsic photoacoustic signals confinement ensures the high lateral and axial resolution, all together. Starting from scratch, I developed and fully studied a loss modulation technique to realize this two-photon acoustic microscopy, investigated potential contrast agents, characterized spatial resolution, enhanced imaging speed by integration with a commercial scanning microscope, and demonstrated biological applications. 0.52 μm lateral and 1.94 μm axial resolution is achieved with high imaging rate, 0.38fps (512×512 pixels). Finally, I utilized this two-photon acoustic microscopy to demonstrate label-free imaging of non-fluorescent melanin distribution within a black mouse ear both ex-vivo and in-vivo.
Subjects
Photoacoustic microscopy
Ti:sapphire laser
two-photon absorption
optical modulation
sub-micron resolution
in-vivo imaging
melanin distribution
Type
thesis
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