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New methods to improve super-resolution imaging techniques give a sharper view inside cells

A sharper view inside cells
Improving axial resolution in 3D SIM. a鈥揷, Schematic representations of beam illumination at objective back focal plane (BFP) and sample planes for wide-field microscopy (single-beam illumination, a), 3D SIM (three-beam illumination, b) and four-beam SIM (a mirror opposite the sample is used to back-reflect the central beam, producing four-beam interference, c). Higher magnification illumination views at right show fine axial structure in four-beam SIM pattern, absent in 3D SIM or wide-field microscopy. d, Axial cross-sectional views of 100-nm beads, as imaged in wide-field microscopy (top), 3D SIM (middle) and four-beam SIM (bottom). e, Higher magnification views of bead highlighted by colored arrowheads in d, illustrating progressive improvement in axial resolution. Insets show magnitude of OTFs (kx/kz plane) derived from images. f, Line profiles corresponding to bead images shown in e, taken along vertical green line in e. g, Quantification of lateral (blue) and axial (orange) FWHM for n鈥=鈥102, 100 and 99 beads for wide-field microscopy, 3D SIM and four-beam SIM, respectively. See also Supplementary Table 1. Whiskers: maximum and minimum; center lines: medians; bounds of box: 75th and 25th percentiles; cross symbols: mean markers. Scale bars, 2鈥壜祄 (d) and 500鈥塶m (e); 1/200鈥塶m鈭1 for Fourier transform insets in e. a.u., arbitrary units. Credit: Nature Biotechnology (2023). DOI: 10.1038/s41587-022-01651-1

New methods to improve super-resolution imaging techniques are giving biologists a clearer and more complete view of the inner workings of living cells.

A new paper in Nature Biotechnology from the Shroff Lab at Howard Hughes Medical Institute (HHMI)'s Janelia Research Campus details two practical ways to improve the axial (or z) resolution of 3D structured illumination microscopy, a technique to see inside living pioneered by former Janelia Group Leader Mats Gustafsson, who died in 2011.

In 3D-SIM and other fluorescence microscopy techniques, the axial resolution of the image is often blurred. This means researchers can clearly see details in two dimensions, on the x and y planes, but details in the third dimension, on the z plane, are fuzzy. Previous attempts to resolve this issue were difficult to implement.

A project led by Xuesong Li, a postdoc in the Shroff Lab, developed two ways to practically deal with the problem. In one method, a mirror is added to the to create an additional beam of light, changing the , and enabling finer, sharper resolution along the z axis.

The second method, which uses , blurs the sharp x and y axes to look like the blurry z axis and then trains a to reverse these blurry images. The network then uses that information to un-blur the z axis.

A new paper in Nature Biotechnology from the Shroff Lab details two practical ways to improve the axial, or z, resolution of 3D structured illumination microscopy, a technique to see inside living cells. In one method, a mirror is added to the microscope to create an additional beam of light, changing the interference pattern, and enabling finer, sharper resolution along the z axis. (video 1). The second method, which uses deep learning, blurs the sharp x and y axes to look like the blurry z axis and then trains a neural network to reverse these blurry images. The network then uses that information to un-blur the z axis. (video 2). Credit: Li et al.

Both methods provide a practical and effective way to improve axial in 3D-SIM. This enables scientists to view the organelles inside cells sharply in all directions and potentially uncover new insights about the components inside cells.

The new methods can be used to improve other microscopes, and the team is working to apply the methods to thicker samples. Some of the same concepts may also be useful in imaging techniques they are developing for use by biologists working in Janelia's new research area, 4D Cellular 麻豆淫院iology.

More information: Xuesong Li et al, Three-dimensional structured illumination microscopy with enhanced axial resolution, Nature Biotechnology (2023).

Journal information: Nature Biotechnology

Citation: New methods to improve super-resolution imaging techniques give a sharper view inside cells (2023, January 26) retrieved 1 May 2025 from /news/2023-01-methods-super-resolution-imaging-techniques-sharper.html
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