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Research presents 2D dipole orientation method for mapping cells

2D dipole orientation method for mapping cells
Schematic illustration of the 3DOM microscopy. Credit: PhotoniX (2024). DOI: 10.1186/s43074-024-00127-6

Due to the high transparency of cells, it is very difficult to observe the organelles within them. Biologists can label specific organelles for observation through fluorescence staining. This is somewhat analogous to being in an environment without light where everyone is dressed entirely in black, making it difficult to find your friends. By having our friends hold a fluorescent stick, we can easily locate them.

An interesting question is: If the angle of the fluorescent stick held by my friend represents a kind of signal, how can we detect such angular information?

Just like this puzzle, due to the highly transparent nature of cells, it is very difficult to observe the organelles in them. With fluorescent staining, biologists can label specific organelles for observation. Most fluorescent molecules appear as directional dipoles during absorption or emission.

The orientation of fluorophores can reveal important information about the structure and dynamics of their associated organelles. Fluorescence polarization microscopy has also developed as an indispensable tool for studying the orientation characteristics of biomolecules.

To overcome the challenge of conventional fluorescence polarization microscopy limited by optical diffraction, improved super-resolution fluorescence polarization microscopy techniques have been proposed, such as single-molecule orientation-localization microscopy (SMOLM) and polarization modulation (e.g., SDOM, SPoD, etc.).

However, from the biotechnological point of view, despite the significant role of biological filaments (e.g., actin filaments and microtubules) in cellular functions, there is a lack of approaches with 3D orientation resolving and high temporal-spatial resolution to study them in vivo.

2D dipole orientation method for mapping cells
Credit: PhotoniX (2024). DOI: 10.1186/s43074-024-00127-6

To address the problem of dipole orientation resolution, Professor Xi Peng's research group from Peking University has developed a 2D dipole orientation mapping method, SDOM, and optical lock-in detection super-resolution dipole orientation mapping, OLID-SDOM. In PhotoniX, the research group report a super-resolution 3D orientation mapping microscope termed 3DOM.

The 3DOM method is based on the polarized structured illumination microscopy developed by the research group. Reversing the principle of Young's double-slit interference and combining it with the principle of reversible light paths, different angles of the stripes are used to produce positive and negative first-order beams in different directions.

Furthermore, a single direction of tilted illumination can be produced by simply blocking the corresponding negative first-order light. By projecting this tilt to different angles of the z-axis and reconstructing the image using the FISTA algorithm, high-precision resolution of the dipole orientation can be achieved by combining the polarization modulation coefficients and the reconstruction results in reciprocal space.

Overall, the proposed 3DOM method effectively overcomes the limitations of fluorescence polarization microscopy in spatial resolution and 3D orientation mapping using widefield imaging.

3DOM provides a more comprehensive understanding of the 3D spatial structure of fluorophore molecules. This enables us not only to distinguish various cytoskeletal organizations ( and microtubules) but also to gain valuable insights into filament binding compactness and the order of subcellular structures.

Moreover, 3DOM holds significant potential in DNA bending and the orientation of membranous organelles. One of the key advantages of 3DOM is its ease of upgradability to existing widefield systems. The simple implementation, accurate 3D dipole orientation information, and superior spatiotemporal resolution of 3DOM make it suitable for a wide range of applications, enhancing its accessibility and usability in different research settings.

This powerful tool empowers researchers to unravel the intricate complexities of subcellular structure, biomechanics, and biodynamics, revolutionizing our understanding of cellular processes. The researchers foresee 3DOM advancing understanding across a multitude of biological structures and interactions operative at the nanoscale.

More information: Suyi Zhong et al, Three-dimensional dipole orientation mapping with high temporal-spatial resolution using polarization modulation, PhotoniX (2024). DOI: 10.1186/s43074-024-00127-6

Provided by Peking University

Citation: Research presents 2D dipole orientation method for mapping cells (2024, May 9) retrieved 14 June 2024 from
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