Intracytoplasmic sperm injection (ICSI) is the most common practice for in vitro fertilization (IVF) treatments. In ICSI, a single sperm is selected and injected into an oocyte. The quality of the sperm and specifically its DNA fragmentation index (DFI) have significant effects on the fertilization success rate. In our research, we use computer vision and deep learning methods to predict DFI scoring for a single sperm cell. Each cell in the dataset was acquired using multiple white light microscopy techniques combined with state-of-the-art interferometry. In our results, we see a strong correlation between the stained images and our score prediction which can be used in the ICSI process.
Typically, tomographic reconstruction of dynamic 3D samples is limited by sample movement speed and illumination scanning speed. However, using the six-pack holography technique that we have developed, 6 off-axis holograms of different illumination angles can be acquired in a single camera exposure, without sacrificing individual image resolution or field of view, and without requiring sample sparsity. Using these 6 holograms we reconstruct the 3D refractive index tomogram of a dynamic sample from each video frame using the optical diffraction tomography algorithm. We demonstrate our technique experimentally on flowing microbeads and swimming microalgae.
We report a module employing shearing interferometry with constant off-axis angle (SICA) that works both for coherent light sources and low-coherence light sources, in which the shearing distance in uncoupled to the off-axis angle. In the module, two laterally sheared sample beams are generated by a grating, and the part of the sample area containing no sample details serves as the reference beam, as in any shearing interferometer. However, in our module, the shearing distance can be flexibly adjusted by controlling the axial location of the grating, while the off-axis interference angle remains constant.
Six-pack holography may be used to reject out-of-focus objects in dynamic samples. By illuminating the sample from six different angles in parallel, out-of-focus objects are laterally shifted in six different directions when projected onto the focal plane. Pixel-wise averaging of the six reconstructed images then reduces the impact of out-of-focus objects. The averaged images reduce the contribution of out-of-focus objects by up to 83%, depending on the out-of-focus object axial distance from the focused plane. Simulation results are shown, as well as experimental results for swimming microalgae and flowing microbeads, including numerical refocusing by Fresnel propagation.
Six-pack holography (6PH) is the first holographic imaging modality that is more spatial bandwidth efficient than on-axis holography. 6PH utilizes a low-coherence light source and a phase delay plate to enable the spatial multiplexing of six off-axis holograms without cross-talk between the six sample and six reference beams, and is thus capable of acquiring six off-axis holograms in a single camera exposure. We applied 6PH to synthetic aperture (SA) superresolution, and produced an SA capable of dynamically increasing the resolution by a factor of 1.62. 6PH presents a valuable general purpose technique for all holography applications and label-free imaging.
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