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Evaluation of Oolong Tea Extract Staining of Brain Tissue with Special Reference to Smooth Endoplasmic Reticulum

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Abstract

Electron microscopy remains the gold standard for studying the nervous system, as it allows adequate spatial resolution imaging of the finer structures of nervous tissues. Despite the high resolution, correct interpretation of electron microscopy images is only possible with sufficient contrast of unit membranes, cytosolic components and the cytoskeleton, which is achieved using hazardous reagents and heavy metals. Earlier, a solution of dry oolong tea extract was introduced as a replacement for toxic and radioactive uranyl acetate. However, staining with oolong extract showed less contrast of intracellular organelles. It is known that depending on the specimen nature and the experimental conditions, different tissues have unequal ability to capture the stain. Here, oolong tea extract staining of early postnatal and adult rat brain tissue was used for the first time to evaluate it with transmission electron microscopy. We found that oolong extract obtained from freshly prepared tea is equal to conventional stains in section staining and has several advantages. We showed that its use on the immature brain allows the finest details of the smooth endoplasmic reticulum of neurons to be revealed. Also, in combination with osmication with potassium ferricyanide and cacodylate buffer, it provides better visualization of the neuronal cytoskeleton and the smooth cisternae in the terminal astrocytic lamellae. A conventional chemical fixation protocol followed by staining with a saturated oolong tea water extract is also well suited for routine studies of the brain of adult animals, as well as for multiple grids batch staining.

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ACKNOWLEDGMENTS

The authors are deeply grateful to Vera G. Tsyganova (ITEB RAS) for her help in handling with neonatal brain and hippocampal material provided. The study was performed at Electron Microscopy Core Facilities of the PSCBR RAS (https://www.pbcras.ru/services/tskp/; http://www.ckp-rf.ru/ckp/670266/), Pushchino, Russia and in Electron Microscopy Suite of the Open University, Milton Keynes, UK.

Funding

This study was conducted in the framework of the State assignment of PSCBR RAS, project number АААА-А20-120101390069-4, and supported by RFBR, project number 20-34-90068.

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Authors and Affiliations

  1. Institute of Cell Biophysics, Russian Academy of Sciences, 142290, Pushchino, Moscow oblast, Russia

    E. A. Shishkova & V. V. Rogachevsky

  2. Faculty of Health and Social Care, the Open University, Walton Hall, MK7 6AA, Milton Keynes, UK

    I. V. Kraev

Authors
  1. E. A. Shishkova
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  2. I. V. Kraev
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  3. V. V. Rogachevsky
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Corresponding author

Correspondence to V. V. Rogachevsky.

Additional information

Abbreviations: SEM, scanning electron microscopy; OTO, osmium–thiocarbohydrazide–osmium; PSD, postsynaptic density; TEM, transmission-electron microscopy; UAc, uranyl acetate; OTE, oolong tea extract; sER, smooth endoplasmic reticulum; UL, dye based on lanthanide salts (UranyLess).

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Shishkova, E.A., Kraev, I.V. & Rogachevsky, V.V. Evaluation of Oolong Tea Extract Staining of Brain Tissue with Special Reference to Smooth Endoplasmic Reticulum. BIOPHYSICS 67, 752–760 (2022). https://doi.org/10.1134/S0006350922050177

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  • DOI: https://doi.org/10.1134/S0006350922050177

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