Label-free superior contrast with c-band ultra-violet extinction microscopy

Florian Ströhl; Deanna L. Wolfson; Ida S. Opstad; Daniel H. Hansen; Hong Mao; Balpreet S. Ahluwalia

doi:10.1038/s41377-023-01105-6

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Label-free superior contrast with c-band ultra-violet extinction microscopy

Original Articles

- Label-free superior contrast with c-band ultra-violet extinction microscopy
- Label-free superior contrast with c-band ultra-violet extinction microscopy
- 光：科学与应用 2023年12卷第3期页码：435-445
- Affiliations：
  
  1.Department of Physics and Technology, UiT The Arctic University of Norway, Tromsø, Norway
  2.Department of Clinical Science, Intervention and Technology, Karolinska Institute, Stockholm, Sweden
- Author bio：
  
  Florian Ströhl (florian.strohl@uit.no)
- Funds：
- DOI：10.1038/s41377-023-01105-6
  中图分类号：
- 纸质出版日期：2023-3-31，
  
  网络出版日期：2023-3-3，
  
  收稿日期：2022-10-27，
  
  修回日期：2023-2-14，
  
  录用日期：2023-2-14
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Label-free superior contrast with c-band ultra-violet extinction microscopy[J]. 光：科学与应用, 2023,12(3):435-445.

Ströhl, F. et al. Label-free superior contrast with c-band ultra-violet extinction microscopy. Light: Science & Applications, 12, 435-445 (2023).
Label-free superior contrast with c-band ultra-violet extinction microscopy[J]. 光：科学与应用, 2023,12(3):435-445. DOI： 10.1038/s41377-023-01105-6.

Ströhl, F. et al. Label-free superior contrast with c-band ultra-violet extinction microscopy. Light: Science & Applications, 12, 435-445 (2023). DOI： 10.1038/s41377-023-01105-6.

摘要

Abstract

In 1934

Frits Zernike demonstrated that it is possible to exploit the sample's refractive index to obtain superior contrast images of biological cells. The refractive index contrast of a cell surrounded by media yields a change in the phase and intensity of the transmitted light wave. This change can be due to either scattering or absorption caused by the sample. Most cells are transparent at visible wavelengths

which means the imaginary component of their complex refractive index

also known as extinction coefficient

is close to zero. Here

we explore the use of c-band ultra-violet (UVC) light for high-contrast high-resolution label-free microscopy

as k is naturally substantially higher in the UVC than at visible wavelengths. Using differential phase contrast illumination and associated processing

we achieve a 7- to 300-fold improvement in contrast compared to visible-wavelength and UVA differential interference contrast microscopy or holotomography

and quantify the extinction coefficient distribution within liver sinusoidal endothelial cells. With a resolution down to 215 nm

we are

for the first time in a far-field label-free method

able to image individual fenestrations within their sieve plates which normally requires electron or fluorescence superresolution microscopy. UVC illumination also matches the excitation peak of intrinsically fluorescent proteins and amino acids and thus allows us to utilize autofluorescence as an independent imaging modality on the same setup.

关键词

Keywords

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