Volumetric trans-scale imaging of massive quantity of heterogeneous cell populations in centimeter-wide tissue and embryo

  1. Taro Ichimura
  2. Taishi Kakizuka
  3. Yuki Sato
  4. Keiko Itano
  5. Kaoru Seiriki
  6. Hitoshi Hashimoto
  7. Hiroya Itoga
  8. Shuichi Onami
  9. Takeharu Nagai

  • Curated by eLife

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    eLife assessment

    The important study established a large-scale objective and integrated multiple optical microscopy systems to demonstrate their potential for long-term imaging of the developmental process. The convincing imaging data cover a wide range of biological applications, such as organoids, mouse brains, and quail embryos, but enhancing image quality can further enhance the method's effectiveness. This work will appeal to biologists and imaging technologists focused on long-term imaging of large fields.

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Abstract

We established a volumetric trans-scale imaging system with an ultra-large field-of-view (FOV) that enables simultaneous observation of millions of cellular dynamics in centimeter-wide three-dimensional (3D) tissues and embryos. Using a custom-made giant lens system with a magnification of 2× and a numerical aperture (NA) of 0.25, and a CMOS camera with more than 100 megapixels, we built a trans-scale scope AMATERAS-2, and realized fluorescence imaging with a lateral spatial resolution of approximately 1.2 µm across an FOV of approximately 1.5 × 1.0 cm 2 . The 3D resolving capability was realized through a combination of optical and computational sectioning techniques tailored for our low-power imaging system. We applied the imaging technique to 1.2 cm-wide section of mouse brain, and successfully observed various regions of the brain with sub-cellular resolution in a single FOV. We also performed time-lapse imaging of a 1-cm-wide vascular network during quail embryo development for over 24 hours, visualizing the movement of over 4.0 × 10 5 vascular endothelial cells and quantitatively analyzing their dynamics. Our results demonstrate the potential of this technique in accelerating production of comprehensive reference maps of all cells in organisms and tissues, which contributes to understanding developmental processes, brain functions, and pathogenesis of disease, as well as high-throughput quality check of tissues used for transplantation medicine.

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  1. Version published to 10.7554/elife.93633.1 on eLife
  2. Version published to 10.7554/elife.93633 on eLife
  3. eLife

    Author Response

    We would like to thank the editors and reviewers who took their valuable time to evaluate the manuscript from various perspectives. We are delighted that our technique was found appealing to biologists and imaging technologists. However, we received several comments that the principles and effectiveness of our techniques are often vague and difficult to understand. They also pointed out that the explanations and representations for several figures were not appropriate. We will revise the manuscript to address these issues and make the manuscript more clear and rigorous.

  4. eLife

    eLife assessment

    The important study established a large-scale objective and integrated multiple optical microscopy systems to demonstrate their potential for long-term imaging of the developmental process. The convincing imaging data cover a wide range of biological applications, such as organoids, mouse brains, and quail embryos, but enhancing image quality can further enhance the method's effectiveness. This work will appeal to biologists and imaging technologists focused on long-term imaging of large fields.

  5. eLife

    Reviewer #1 (Public Review):

    Summary:
    The authors are trying to develop a microscopy system that generates data output exceeding the previous systems based on huge objectives.

    Strengths:
    They have accomplished building such a system, with a field of view of 1.5x1.0 cm2 and a resolution of up to 1.2 um. They have also demonstrated their system performance on samples such as organoids, brain sections, and embryos.

    Weaknesses:
    To be used as a volumetric imaging technique, the authors only showcase the implementation of multi-focal confocal sectioning. On the other hand, most of the real biological samples were acquired under wide-field illumination, and processed with so-called computational sectioning. Despite the claim that it improves the contrast, sometimes I felt that the images were oversharpened and the quantitative nature of these fluorescence images may be perturbed.

  6. eLife

    Reviewer #2 (Public Review):

    Summary:
    This manuscript introduced a volumetric trans-scale imaging system with an ultra-large field-of-view (FOV) that enables simultaneous observation of millions of cellular dynamics in centimeter-wide 3D tissues and embryos. In terms of technique, this paper is just a minor improvement of the authors' previous work, which is a fluorescence imaging system working at visible wavelength region (https://www.nature.com/articles/s41598-021-95930-7).

    Strengths:
    In this study, the authors enhanced the system's resolution and sensitivity by increasing the numerical aperture (NA) of the lens. Furthermore, they achieved volumetric imaging by integrating optical sectioning and computational sectioning. This study encompasses a broad range of biological applications, including imaging and analysis of organoids, mouse brains, and quail embryos, respectively. Overall, this method is useful and versatile.

    Weaknesses:
    The unique application that only can be done by this high-throughput system remains vague. Meanwhile, there are also several outstanding issues in this paper, such as the lack of technical advances, unclear method details, and non-standardized figures.

  7. Version published to 10.1101/2023.08.21.553997v2 on bioRxiv
  8. Version published to 10.1101/2023.08.21.553997v1 on bioRxiv