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Digital Holographic Microscopy (DHM) is a potentially non-invasive new technology which can be applied in many
areas from applied imaging science to biomedical optics. DHM is an interferometric technique that gives us a number of
important advantages such as the possibility to acquire holograms at high speed, to obtain complete information about
amplitude and phase and to use image processing techniques. In this sense, DHM offers rapid 3D imaging with a
theoretically higher resolution than OCT (Optical Coherent Tomography). By this technique optical path measurements
with sensitivities in the nanometer range of reflective and transparent objects can be obtained.
In this work, we use DHM to study the effect of ablation using 4.5 nJ pulses on chicken corneas. For this, a titanium
sapphire laser at 800 nm and 76 MHz frequency (Vitesse, Coherent Inc. USA) was focused to its diffraction-limited spot
size by a 10x objective of 0.3 numerical aperture. The width of the pulse (170 fs) at the sample was measured by spectral
techniques. The average beam power at the sample was 340 mW and all the system was mechanically driven by a XY
synchronization unit that controls the speed of the sample movement. The speed of the sample was varied between 1-50
μm/s.
The studied chicken corneal tissue was previously processed by Trypan dye in order to visualize the irradiated area. The
photodisrupted zone was analyzed by a HDM technique by illuminating it using a laser diode source (λ=683 nm) linearly
polarized in a modified Mach-Zehnder with an off-axis geometry configuration. The reflected object wave by the tissue
surface (specimen) interferes with the reference wave and a CCD camera records the hologram. As a result, the influence
of the speed of photodisruption in the depth of the ablated corneas was analyzed. Therefore, it is possible to analyze
thermal and photoirradiated effects on corneal tissues which allow us the possibility to optimize the interaction of
intratissue and the intratissue target region of interest.
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