1. Cell Biology

A versatile Oblique Plane Microscope for large-scale and high-resolution imaging of subcellular dynamics

  1. Etai Sapoznik
  2. Bo-Jui Chang
  3. Jaewon Huh
  4. Robert J Ju
  5. Evgenia V Azarova
  6. Theresa Pohlkamp
  7. Erik S Welf
  8. David Broadbent
  9. Alexandre F Carisey
  10. Samantha J Stehbens
  11. Kyung-Min Lee
  12. Arnoldo Marin
  13. Ariella B Hanker
  14. Jens C Schmidt
  15. Carlos L Arteaga
  16. Bin Yang
  17. Yoshihiko Kobayashi
  18. Purushothama Rao Tata
  19. Rory Kruithoff
  20. Konstantin Dubrovinski
  21. Doug P Shepherd
  22. Alfred Millet-Sikking
  23. Andrew G York
  24. Kevin M Dean  Is a corresponding author
  25. Reto P Fiolka  Is a corresponding author
  1. University of Texas Southwestern Medical Center, United States
  2. University of Queensland, Australia
  3. Michigan State University, United States
  4. Baylor College of Medicine and Texas Children's Hospital, United States
  5. Chan Zuckerberg Biohub, United States
  6. Duke University, United States
  7. University of Arizona, United States
  8. Calico Life Sciences LLC, United States
https://doi.org/10.7554/eLife.57681
Share this article
Cite this article
  1. Etai Sapoznik
  2. Bo-Jui Chang
  3. Jaewon Huh
  4. Robert J Ju
  5. Evgenia V Azarova
  6. Theresa Pohlkamp
  7. Erik S Welf
  8. David Broadbent
  9. Alexandre F Carisey
  10. Samantha J Stehbens
  11. Kyung-Min Lee
  12. Arnoldo Marin
  13. Ariella B Hanker
  14. Jens C Schmidt
  15. Carlos L Arteaga
  16. Bin Yang
  17. Yoshihiko Kobayashi
  18. Purushothama Rao Tata
  19. Rory Kruithoff
  20. Konstantin Dubrovinski
  21. Doug P Shepherd
  22. Alfred Millet-Sikking
  23. Andrew G York
  24. Kevin M Dean
  25. Reto P Fiolka
(2020)
A versatile Oblique Plane Microscope for large-scale and high-resolution imaging of subcellular dynamics
eLife 9:e57681.
https://doi.org/10.7554/eLife.57681
  2. Download BibTeX
  3. Download .RIS

Abstract

We present an Oblique Plane Microscope that uses a bespoke glass-tipped tertiary objective to improve the resolution, field of view, and usability over previous variants. Owing to its high numerical aperture optics, this microscope achieves lateral and axial resolutions that are comparable to the square illumination mode of Lattice Light-Sheet Microscopy, but in a user friendly and versatile format. Given this performance, we demonstrate high-resolution imaging of clathrin-mediated endocytosis, vimentin, the endoplasmic reticulum, membrane dynamics, and Natural Killer-mediated cytotoxicity. Furthermore, we image biological phenomena that would be otherwise challenging or impossible to perform in a traditional light-sheet microscope geometry, including cell migration through confined spaces within a microfluidic device, subcellular photoactivation of Rac1, diffusion of cytoplasmic rheological tracers at a volumetric rate of 14 Hz, and large field of view imaging of neurons, developing embryos, and centimeter-scale tissue sections.

Data availability

Manuscript data is available on Zenodo, under the doi:10.5281/zenodo.4266823.

The following data sets were generated

Article and author information

Author details

  1. Etai Sapoznik

    Cell Biology, University of Texas Southwestern Medical Center, Dallas, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8472-0299

  2. Bo-Jui Chang

    Cell Biology, University of Texas Southwestern Medical Center, Dallas, United States
    Competing interests
    No competing interests declared.

  3. Jaewon Huh

    Bioinformatics and Cell Biology, University of Texas Southwestern Medical Center, Dallas, United States
    Competing interests
    No competing interests declared.

  4. Robert J Ju

    Institute for Molecular Bioscience, University of Queensland, St Lucia, Queensland, Australia
    Competing interests
    No competing interests declared.

  5. Evgenia V Azarova

    Department of Cell Biology, University of Texas Southwestern Medical Center, Dallas, United States
    Competing interests
    No competing interests declared.

  6. Theresa Pohlkamp

    Department of Molecular Genetics, University of Texas Southwestern Medical Center, Dallas, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-3923-1917

  7. Erik S Welf

    Cell Biology, University of Texas Southwestern Medical Center, Dallas, United States
    Competing interests
    No competing interests declared.

  8. David Broadbent

    nstitute for Quantitative Health Sciences and Engineering, Michigan State University, East Lansing, United States
    Competing interests
    No competing interests declared.

  9. Alexandre F Carisey

    William T. Shearer Center for Human Immunobiology, Baylor College of Medicine and Texas Children's Hospital, Houston, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1326-2205

  10. Samantha J Stehbens

    Institute for Molecular Bioscience, University of Queensland, St. Lucia, Queensland, Australia
    Competing interests
    No competing interests declared.

  11. Kyung-Min Lee

    Harold C Simmons Comprehensive Cancer Center and the Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, United States
    Competing interests
    No competing interests declared.

  12. Arnoldo Marin

    Harold C Simmons Comprehensive Cancer Center and the Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, United States
    Competing interests
    No competing interests declared.

  13. Ariella B Hanker

    Harold C Simmons Comprehensive Cancer Center and the Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, United States
    Competing interests
    No competing interests declared.

  14. Jens C Schmidt

    OBGYN, Institute for Quantitative Health Sciences and Engineering, Michigan State University, East Lansing, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-9061-7853

  15. Carlos L Arteaga

    Harold C. Simmons Comprehensive Cancer Center and the Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, United States
    Competing interests
    Carlos L Arteaga, C.L.A. serves in an advisory role for Novartis, which has an investment interest in alpelisib..

  16. Bin Yang

    Chan Zuckerberg Biohub, Chan Zuckerberg Biohub, San Francisco, United States
    Competing interests
    No competing interests declared.

  17. Yoshihiko Kobayashi

    Department of Cell Biology, Duke University, Durham, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-7031-1478

  18. Purushothama Rao Tata

    Department of Cell Biology, Duke University, Durham, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-4837-0337

  19. Rory Kruithoff

    Department of Physics and the Center for Biological Physics, University of Arizona, Tempe, United States
    Competing interests
    No competing interests declared.

  20. Konstantin Dubrovinski

    Systems Biology, University of Texas Southwestern Medical Center, Dallas, United States
    Competing interests
    No competing interests declared.

  21. Doug P Shepherd

    Department of Physics and the Center for Biological Physics, University of Arizona, Tempe, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-9087-0832

  22. Alfred Millet-Sikking

    Calico, Calico Life Sciences LLC, South San Franscisco, United States
    Competing interests
    No competing interests declared.

  23. Andrew G York

    Calico, Calico Life Sciences LLC, South San Franscisco, United States
    Competing interests
    Andrew G York, K.M.D. has an investment interest in Discovery Imaging Systems, LLC.

  24. Kevin M Dean

    Cell Biology, University of Texas Southwestern Medical Center, Dallas, United States
    For correspondence
    Kevin.Dean@UTsouthwestern.edu
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-0839-2320

  25. Reto P Fiolka

    Cell Biology, University of Texas Southwestern Medical Center, Dallas, United States
    For correspondence
    Reto.Fiolka@UTsouthwestern.edu
    Competing interests
    Reto P Fiolka, The author has an investment interest in Discovery Imaging Systems, LLC..
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4636-5000

Funding

Cancer Prevention and Research Institute of Texas (RR160057)

  • Reto P Fiolka

National Institutes of Health (5P30CA142543)

  • Kevin M Dean

Damon Runyon Cancer Research Foundation (DFS-24-17)

  • Jens C Schmidt

Chan Zuckerberg Initiative (HCA3-0000000196)

  • Purushothama Rao Tata

Chan Zuckerberg Initiative (HCA3-0000000196)

  • Doug P Shepherd

Chan Zuckerberg Initiative (HCA3-0000000196)

  • Yoshihiko Kobayashi

ARC (FT190100516)

  • Samantha J Stehbens

Rebecca Cooper Medical Foundation (PG2018168)

  • Samantha J Stehbens

University of Queensland Early Career Award (RM2018002613)

  • Samantha J Stehbens

Company of Biologists (JCSTF1903138)

  • Robert J Ju

Robert A. Welch Foundation (I-1950-20180324)

  • Konstantin Dubrovinski

National Institutes of Health (R00 GM120386)

  • Jens C Schmidt

National Institutes of Health (R01GM110066)

  • Konstantin Dubrovinski

National Institutes of Health (R01HL068702)

  • Doug P Shepherd

National Institutes of Health (R33CA235254)

  • Reto P Fiolka

National Institutes of Health (R35GM133522)

  • Reto P Fiolka

National Institutes of Health (K25 CA204526)

  • Erik S Welf

National Institutes of Health (P30 CA142543)

  • Carlos L Arteaga

National Institutes of Health (1R01MH120131-01A1)

  • Kevin M Dean

National Institutes of Health (1R34NS121873)

  • Kevin M Dean

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Melike Lakadamyali, University of Pennsylvania, United States

Version history

  1. Received: April 8, 2020
  2. Accepted: November 9, 2020
  3. Accepted Manuscript published: November 12, 2020 (version 1)
  4. Accepted Manuscript updated: November 16, 2020 (version 2)
  5. Version of Record published: December 1, 2020 (version 3)
  6. Version of Record updated: December 7, 2020 (version 4)
  7. Version of Record updated: February 1, 2021 (version 5)

Copyright

© 2020, Sapoznik et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 30,152
    Page views
  • 1,374
    Downloads
  • 86
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, Scopus, PubMed Central.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Etai Sapoznik
  2. Bo-Jui Chang
  3. Jaewon Huh
  4. Robert J Ju
  5. Evgenia V Azarova
  6. Theresa Pohlkamp
  7. Erik S Welf
  8. David Broadbent
  9. Alexandre F Carisey
  10. Samantha J Stehbens
  11. Kyung-Min Lee
  12. Arnoldo Marin
  13. Ariella B Hanker
  14. Jens C Schmidt
  15. Carlos L Arteaga
  16. Bin Yang
  17. Yoshihiko Kobayashi
  18. Purushothama Rao Tata
  19. Rory Kruithoff
  20. Konstantin Dubrovinski
  21. Doug P Shepherd
  22. Alfred Millet-Sikking
  23. Andrew G York
  24. Kevin M Dean
  25. Reto P Fiolka
(2020)
A versatile Oblique Plane Microscope for large-scale and high-resolution imaging of subcellular dynamics
eLife 9:e57681.
https://doi.org/10.7554/eLife.57681

Share this article

https://doi.org/10.7554/eLife.57681

Categories and tags

Further reading

    1. Cell Biology
    2. Chromosomes and Gene Expression

    Heat stress impairs centromere structure and segregation of meiotic chromosomes in Arabidopsis

    Lucie Crhak Khaitova, Pavlina Mikulkova ... Karel Riha
    Research Article

    Heat stress is a major threat to global crop production, and understanding its impact on plant fertility is crucial for developing climate-resilient crops. Despite the known negative effects of heat stress on plant reproduction, the underlying molecular mechanisms remain poorly understood. Here, we investigated the impact of elevated temperature on centromere structure and chromosome segregation during meiosis in Arabidopsis thaliana. Consistent with previous studies, heat stress leads to a decline in fertility and micronuclei formation in pollen mother cells. Our results reveal that elevated temperature causes a decrease in the amount of centromeric histone and the kinetochore protein BMF1 at meiotic centromeres with increasing temperature. Furthermore, we show that heat stress increases the duration of meiotic divisions and prolongs the activity of the spindle assembly checkpoint during meiosis I, indicating an impaired efficiency of the kinetochore attachments to spindle microtubules. Our analysis of mutants with reduced levels of centromeric histone suggests that weakened centromeres sensitize plants to elevated temperature, resulting in meiotic defects and reduced fertility even at moderate temperatures. These results indicate that the structure and functionality of meiotic centromeres in Arabidopsis are highly sensitive to heat stress, and suggest that centromeres and kinetochores may represent a critical bottleneck in plant adaptation to increasing temperatures.

    1. Cell Biology

    High-altitude hypoxia exposure inhibits erythrophagocytosis by inducing macrophage ferroptosis in the spleen

    Wan-ping Yang, Mei-qi Li ... Qian-qian Luo
    Research Article

    High-altitude polycythemia (HAPC) affects individuals living at high altitudes, characterized by increased red blood cells (RBCs) production in response to hypoxic conditions. The exact mechanisms behind HAPC are not fully understood. We utilized a mouse model exposed to hypobaric hypoxia (HH), replicating the environmental conditions experienced at 6000 m above sea level, coupled with in vitro analysis of primary splenic macrophages under 1% O2 to investigate these mechanisms. Our findings indicate that HH significantly boosts erythropoiesis, leading to erythrocytosis and splenic changes, including initial contraction to splenomegaly over 14 days. A notable decrease in red pulp macrophages (RPMs) in the spleen, essential for RBCs processing, was observed, correlating with increased iron release and signs of ferroptosis. Prolonged exposure to hypoxia further exacerbated these effects, mirrored in human peripheral blood mononuclear cells. Single-cell sequencing showed a marked reduction in macrophage populations, affecting the spleen’s ability to clear RBCs and contributing to splenomegaly. Our findings suggest splenic ferroptosis contributes to decreased RPMs, affecting erythrophagocytosis and potentially fostering continuous RBCs production in HAPC. These insights could guide the development of targeted therapies for HAPC, emphasizing the importance of splenic macrophages in disease pathology.

    1. Cell Biology

    Golgi Apparatus: Making progress

    Jurgen Denecke
    Insight

    Mapping proteins in and associated with the Golgi apparatus reveals how this cellular compartment emerges in budding yeast and progresses over time.