The new microscope enables high-throughput 3D adaptive optical imaging

[Figure 1] (A) Schematic diagram of a compression time inversion matrix microscope. The random speckle field generated by the rotating diffuser sequentially illuminates the sample under the aberrated medium, and the reflected speckle field is measured by an off-axis digital holographic microscope. BS: Beam splitter. OL: Objective lens. (B) An example of a hologram image measured in a speckle field. (Top: Intensity, Bottom: Phase). Credit: DOI: 10.1038 / s41377-021-00705-4

Microscopes are an important tool in biomedical research because they enable detailed observation and imaging of tissues. Due to the opaque nature of biological materials, heavy light scattering occurs as light passes through tissues, resulting in high levels of background noise and complex optical aberrations. Therefore, most light microscopes can see the surface of the tissue, and the depth details of multiple cell layers are out of reach of many microscopes. This makes it very difficult to capture high resolution optical images of fine structures deep in the tissue.

About a year ago, a research group led by Professor Choi Wonshik of the Molecular Spectroscopy and Dynamics Center (CMSD) in the Institute of Fundamental Sciences (IBS) Imaging technology Called “reflection” matrix Microscopy that combines the capabilities of both hardware and advanced adaptive optics of computation. ” In contrast to traditional imaging, it measures a reflection matrix that contains all accessible information about the relationship between the input and output fields of the imaging system, including the object of interest. .. Then, after digital image processing, you can extract a crisp, distortion-free image of the object from the measured matrix.

Thus, this technology has emerged as a good candidate for label-free, non-invasive, high-resolution optical imaging deep in living tissue. Matrix imaging is certainly better than most traditional AOs. For example, researchers have demonstrated that this technique is powerful enough to “see through” the skull of an intact mouse and enable accurate imaging of the underlying neurons.

The new microscope enables high-throughput 3D adaptive optical imaging
[Figure 2] Image reconstruction from a compressed time reversal matrix. (A) An image distorted by a medium that induces aberrations. (B) Aberration-free image restored from a compressed time-reversal matrix. (C) Point image distribution functions before aberration correction (upper left) and after aberration correction (lower left) and their line profiles (right). Credit: DOI: 10.1038 / s41377-021-00705-4

Despite its amazing performance, reflection matrix microscopy was not without its drawbacks. Measuring the entire reflection matrix is ​​time consuming and vulnerable to external perturbations. This is because it is necessary to measure a large number of interfering images of all accessible input lighting fields. More sparse sampling can speed up the process, but inadequate sampling can limit the ability to correct distortion. Therefore, this meant that real-time volumetric imaging of living samples was not possible, putting practical limitations on its application to biodynamic agriculture research.

With the latest research published in the journal Light: Science and application, The same IBS Group recently announced a new and improved version of its previous AO microscopy technology. This new real-time volume AO imaging system enables 3D imaging over a wide depth range on highly aberrated samples while minimizing image degradation.

To speed up data acquisition, Hee-seop Choi’s team used compressed sensing technology in the context of matrix imaging. They introduced a rotating optical diffuser into a previously deployed reflection matrix microscope to sequentially illuminate an unknown speckle pattern on a sample. We then obtained a compressed reflection matrix by taking far fewer speckle images than previously needed. This has reduced the matrix acquisition time by almost 100 times.

The new microscope enables high-throughput 3D adaptive optical imaging
[Figure 3] Relationship between image quality and compression rate (CR). Restored images of 50, 10, and 2 percent CR (top row) and aberration maps (bottom row). Credit: DOI: 10.1038 / s41377-021-00705-4

Image post-processing used a compressed “time inversion matrix” and a proprietary algorithm to identify sample information and aberrations separately. The advantage of this technique is that it not only significantly reduces the matrix acquisition time, but also eliminates the need for careful calibration and specific selection of lighting patterns to use.

Compressed time-reversed matrix imaging enables near real-time volumetric AO imaging. The function of the new microscope was demonstrated by aberration-free 3D imaging of myelin nerve fibers in the mouse brain. Data acquisition time for 128 x 128 x 125 μm volume imaging was only 3.58 seconds3The lateral resolution of the structure and diffraction limit is 0.45 μm, and the axial resolution is 2 μm.

The new microscope enables high-throughput 3D adaptive optical imaging
[Figure 4] Aberration-free volumetric image of the mouse brain taken by a compression-time inversion matrix microscope. (A) Imaging configuration. 3D imaging of the mouse brain continuously illuminates an uncontrolled dynamic speckle pattern and captures a small number of speckle images while moving vertically through the sample stage. (B) A 3D image of a mouse brain reconstructed from a compressed time-reversed matrix. (C) Pupil aberration map corresponding to typical cross-sectional images before and after aberration correction. (D) Line profile of myelinated fibers before and after aberration correction. Credit: DOI: 10.1038 / s41377-021-00705-4

It is expected to open new avenues for the practical application of matrix imaging in all fields of wave engineering, including biomedical imaging. Choi said, “Faster reflection matrix imaging technology is expected to enable real-time non-destructive 3D optical diagnosis in the future, leading to faster diagnosis and progress in neuroscience research. Applications in all fields of wave engineering, including. ”

Scientists have invented a new type of microscope that can be seen through an intact skull

For more information:
Hojun Lee et al, High Throughput Volume Adaptive Optical Imaging with Compressed Time Inversion Matrix, Light: Science and application (2022). DOI: 10.1038 / s41377-021-00705-4

Quote: The new microscope is a high-throughput 3D adaptive optical imaging (2022,) acquired on February 14, 2022 from https: // February 14th) will be possible. html

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The new microscope enables high-throughput 3D adaptive optical imaging

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