Digital Holographic Microscopy (DHM) has become a powerful diagnostics tool for the sciences, especially biology. In addition, DHM has great potential for use in education because it is so flexible and affordable to schools and students. For many years, the cost and complexity of producing and using holograms limited their use to a narrow range of industry and science. Digital holography has removed these limitations making holography affordable and available to anyone who has a laptop computer. Affordable digital holocameras entering the market can enable teachers and students to produce and view their own holograms, reconstruct and view three dimensional images, and better understand and use holography in research. This paper describes such a system including exercises and experiments that explain how holograms are made and how they are used to record and analyze dynamic events that take place in three dimensions. MetroLaser’s “Holoscope”, is a lensless, DHM that produces holographic videos of microscopic objects distributed in a relatively large volume. Each frame in the video is a digital hologram that encodes 3D images that can be focused and viewed, plane by plane in detail. The three-dimensional images encoded in the holograms are electronically reconstructed, scanned, and viewed with sharp images coming in and out of focus on a computer monitor as we scan through the volume. By providing micrometer resolution throughout a cubic centimeter volume, the system effectively freezes time at the moment each frame was recorded and enables precisely tracking the 3D movement of microscopic objects in space and time. The ability to view each hologram in real-time, shows directly how interference between an object and reference wave produces interference fringes that become the hologram. Software provided with the system enables the viewer to automatically perform all of the necessary operations from hologram recording to 3D wavefront reconstruction and plane by plane imaging. We also describe more advanced and more expensive DHMs with even higher resolution that are now also available for scientific research.
This work presents a versatile digital holography software (HOLO4D) that provides an all-in-one solution for holographic reconstruction, simulation, de-twining and lens distortion removal in a user-friendly graphical interface. The software accepts holograms for both, in-line and off-axes schemes with the option to locate and specify the cross-term in the latter case. Both amplitude and phase shifts are recovered from holograms generated by plane waves using multiple functionalities for propagation. These include the angular spectrum method, sinc-interpolation, upsampling-lowpass filter and zero-padding processes that can be applied in a variety of combinations to optimize holographic reconstructions. The virtual image that obscures the real image in these reconstructions is suppressed using a subtraction division plus mean (SDPM) normalization procedure or by iteratively updating the complex-valued wavefront in the detector plane. The use of lenses to manipulate the effective position of the hologram introduces optical distortions. These are removed by calibrating a dot-pattern hologram reconstruction in a step-by-step auto-detection procedure and mapping the distortion characteristics to the corresponding object hologram reconstruction. Hologram simulations enable determining the maximum field-of-view, fringes captured, and the lateral resolution possible based on the imaging sensor, illumination and setup specifications. The numerical package is used to process holograms of 80 μm thick hair strands, scan the depth-of-field and locate the precise three-dimensional (3D) location of reconstructed objects. The results demonstrate the versatility of the software to reconstruct, de-twin, undistort and simulate holograms in a user friendly manner and its applicability as a robust tool for 3D visualization, education, and holographic research.
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