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Cryoconveyor Protocols in Correlative Light and Electron Microscopy: From Multilevel Imaging to Modeling the Biophysical Effects and “Cryotheranostics”

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Abstract

This paper is a technical and methodological note aimed to introduce into the practice of biological research methods a cryomicroscopy in a conveyor mode from small magnifications to the limits of magnification/resolution of scanning electron cryomicroscopy (CryoSEM). The protocol described can be applied to the samples of a low sample preparation complexity without ultratomy or processing typical of transmission electron microscopy. According to this protocol, samples are analyzed in a single microcuvette (chip) indexed by laboratory information management system and sequentially transferred from the nondestructive low-resolution optical microscopy instruments, such as lensless cryo-microscopes, to the CryoSEM/CryoESEM level (in programmable environments and atmospheres). Methods that were introduced and tested include correlative lensless cryomicroscopy and CryoSEM (including those with the sequential transition to microanalysis on wavelength-dispersive X-ray spectrometer on the Rowland circle) as well as microscopy and microinterferometry methods in the ranges from infrared to far-ultraviolet. Among the advantages of the cryoconveyor analysis protocols are sample preservation in a single portable cuvette-chip along with a possibility to establish spatial colocalization between data of optical and electron microscopy using pattern recognition software (all the way to indexing in the laboratory information system) in conducting a full range of microscopic examination. An opportunity is additionally provided for a comprehensive nondestructive sample analysis in a sequential study of microscopic systems with a possibility of variation at the subsequent stages of high-resolution microscopy, depending on the results obtained at the preceding stages of microscopy (at lower resolution).

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ACKNOWLEDGMENTS

I thank my colleagues and students from Moscow State University, Semenov Federal Research Center for Chemical Physics, and Emanuel Institute of Biochemical Physics of the Russian Academy of Sciences for their participation in practical evaluation of the correlative microscopy and cryomicroscopy methods.

Funding

The research was carried out at the sole cost and expense of the author. No additional grants to carry out or direct this particular research were obtained.

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Authors and Affiliations

  1. Semenov Federal Research Center for Chemical Physics, Russian Academy of Sciences, 119991, Moscow, Russia

    O. V. Gradov

  2. Institute of Biochemical Physics, Russian Academy of Sciences, 119334, Moscow, Russia

    O. V. Gradov

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  1. O. V. Gradov
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Correspondence to O. V. Gradov.

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Translated by E. Kuznetsova

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Gradov, O.V. Cryoconveyor Protocols in Correlative Light and Electron Microscopy: From Multilevel Imaging to Modeling the Biophysical Effects and “Cryotheranostics”. Moscow Univ. Biol.Sci. Bull. 78 (Suppl 1), S64–S68 (2023). https://doi.org/10.3103/S0096392523700244

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