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<p>Cryogenic electron microscopy is a powerful tool in structural biology. In thick specimens, challenges arise as an exponentially larger fraction of the transmitted electrons lose energy from inelastic scattering and can no longer be properly focused as a result of chromatic aberrations in the post-specimen optics. Rather than filtering out the inelastic scattering at the price of reducing potential signal, as is done in energy-filtered transmission electron microscopy, we show how a dose-efficient and unfiltered image can be rapidly obtained using tilt-corrected bright-field scanning transmission electron microscopy data collected on a pixelated detector. Enhanced contrast and a 3–5× improvement in dose efficiency are observed for two-dimensional images of intact bacterial cells and large organelles using tilt-corrected bright-field scanning transmission electron microscopy compared to energy-filtered transmission electron microscopy for thicknesses beyond 500 nm. As a proof of concept for the technique’s performance in structural determination, we present a single-particle analysis map at sub-nanometer resolution for a highly symmetric virus-like particle determined from 789 particles.</p>

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Dose-efficient cryo-electron microscopy for thick samples using tilt- corrected scanning transmission electron microscopy

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年9月23日

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  • Yue Yu,
  • Katherine A. Spoth,
  • Michael Colletta,
  • Kayla X. Nguyen,
  • Steven E. Zeltmann,
  • Xiyue S. Zhang,
  • Mohammadreza Paraan,
  • Mykhailo Kopylov,
  • Charlie Dubbeldam,
  • Daniel Serwas,
  • Hannah Siems,
  • David A. Muller,
  • Lena F. Kourkoutis

摘要

Cryogenic electron microscopy is a powerful tool in structural biology. In thick specimens, challenges arise as an exponentially larger fraction of the transmitted electrons lose energy from inelastic scattering and can no longer be properly focused as a result of chromatic aberrations in the post-specimen optics. Rather than filtering out the inelastic scattering at the price of reducing potential signal, as is done in energy-filtered transmission electron microscopy, we show how a dose-efficient and unfiltered image can be rapidly obtained using tilt-corrected bright-field scanning transmission electron microscopy data collected on a pixelated detector. Enhanced contrast and a 3–5× improvement in dose efficiency are observed for two-dimensional images of intact bacterial cells and large organelles using tilt-corrected bright-field scanning transmission electron microscopy compared to energy-filtered transmission electron microscopy for thicknesses beyond 500 nm. As a proof of concept for the technique’s performance in structural determination, we present a single-particle analysis map at sub-nanometer resolution for a highly symmetric virus-like particle determined from 789 particles.