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<p>Correlative light and electron microscopy is a scientific method that encompasses several technologies and workflows. One of the pioneering workflows consisted in vitrifying by high pressure freezing, a sample shortly after live observation (Verkade in J Microsc 230:317–328, 2008). Despite its extraordinary potential, it did not turn into a routine technique for practical reasons. We redesigned the entire tool set, from the CryoCapsule sample carrier to the HPM Live µ high-pressure freezing machine and the R221 embedding resin, to standardize and democratise the technique. In our manuscript, we present all the technological developments that lead to a routine workflow for live to HPF CLEM. We demonstrate our ability to track rapidly moving endosomes live and retrieve them at the electron microscopy level, in three dimensions, with high confidence.</p>

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Combining the CryoCapsule, HPM live µ and R221 conductive resin to track endosome from live imaging to electron microscopy using high-pressure freezing

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年12月4日

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  • Xavier Heiligenstein,
  • Chie Kodera,
  • Yann Bret,
  • Frederic Eyraud,
  • Jérôme Heiligenstein,
  • Martin Belle

摘要

Correlative light and electron microscopy is a scientific method that encompasses several technologies and workflows. One of the pioneering workflows consisted in vitrifying by high pressure freezing, a sample shortly after live observation (Verkade in J Microsc 230:317–328, 2008). Despite its extraordinary potential, it did not turn into a routine technique for practical reasons. We redesigned the entire tool set, from the CryoCapsule sample carrier to the HPM Live µ high-pressure freezing machine and the R221 embedding resin, to standardize and democratise the technique. In our manuscript, we present all the technological developments that lead to a routine workflow for live to HPF CLEM. We demonstrate our ability to track rapidly moving endosomes live and retrieve them at the electron microscopy level, in three dimensions, with high confidence.