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Such a laser would be valuable for extended depth SSOCT imaging applications, and would even further simplify this technique for resolving the complex conjugate ambiguity in SSOCT.
Brightfield images were taken using either a Nikon Eclipse E800 microscope, which allowed for extended depth of field capabilities using Nikon Elements (v. 4) software or with a Nikon Eclipse E400 that used ACT 1 software.
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The system is not designed for optically sectioned imaging and the quality of extended depth fluorescent samples was similar to that seen from a conventional wide field microscope with a similar NA objective lens.
In addition, threshold values were independently determined for each channel by using extended depth of field measurements, in which threshold values were typically found to vary between 10 15 units for each channel from the isosurface values.
The viewsheds examined in this study had large areal extents, large relief, extended depth, and naturalness, and may extend beyond the spatial scales for which view-quality concepts and preference relationships were developed.
At this point, anaerobic cultures were shifted to air and cells were collected at 30, 60 and 120 min. The 488 nm laser was used for excitation and fluorescence images using extended depth of field were collected on channel 3 (560–595 nm), and bright-field images were collected on channels 1 and 9. Between 10 000 and 50 000 cells were acquired for statistical analysis.
Z-stack images spanning the 2 μm from the surface were acquired, underwent 3D deconvolution using AutoQuant (MediaCybernetics, MD, USA) to reduce the distortion and produce a clearer image, and then flattened using Extended Depth of Field for maximum intensity to obtain a single image (ImagePro Plus, MediaCybernetics, MD, USA).
Because accurate, high-throughput phenotypic characterisation is crucial to the success of such endeavours, we recently developed an optical coherence tomography (OCT) system with extended depth range scanning capability for measuring ocular component dimensions in mice.
For the production of images with an extended depth of field, the sequential Z-stack images were taken with 2 μm optical Z sections.
Introducing a second camera, customizing the spectrometer for extended scan depth at the expense of resolution [ 39], or implementing complex conjugate artifact removal [ 40– 43] are alternative customized strategies for imaging the deeper structures required for axis determination.
– 3 megapixel camera, LED Flash, no zoom but extended depth of field.
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