Abstract
Epoxy-embedded semithin sections are useful for the analysis of cell and tissue organization, as well as for the processing of samples for transmission electron microscopy. Because only a very limited number of staining protocols have been developed for epoxy-embedded sections; semithin sections are used infrequently compared to conventional paraffin sections. Here, we describe a simple and reproducible polychromatic protocol for the routine staining of epoxy-embedded semithin sections by adapting Twort’s staining method (mixture of neutral red and fast green FCF). The method can be used for the visualization of cellular organization as well as for the detection of elastic and collagen fibers. The proposed protocol demonstrated the best results for samples fixed for transmission electron microscopy, which suggests, as we demonstrated here, that this staining protocol can also be used for correlative light and electron microscopy.
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References
Aparicio SR, Marsden P (1969) A rapid methylene blue-basic fuchsin stain for semi-thin sections of peripheral nerve and other tissues. J Microsc 89:139–141. https://doi.org/10.1111/j.1365-2818.1969.tb00659.x
Bourne CA, St John DJ (1978) Improved procedure for polychromatic staining of epoxy sections with basic fuchsin–toluidine blue. Med Lab Sci 35:399–400
Collan Y (1969) Staining of epoxy-embedded tissue sections for light microscopy. Exp Path 3:147–152
D’Amico F (2005) A polychromatic staining method for epoxy embedded tissue: a new combination of methylene blue and basic fuchsine for light microscopy. Biotech Histochem 80:207–210. https://doi.org/10.1080/10520290600560897
di Sant’ Agnese PA, De Mesy Jensen KL (1984) Dibasic staining of large epoxy tissue sections and applications to surgical pathology. Am J Clin Pathol 81:25–29. https://doi.org/10.1093/ajcp/81.1.25
Grimley PM (1964) A tribasic stain for thin sections of plastic-embedded, OsO4-fixed tissues. Stain Technol 39:229–233. https://doi.org/10.3109/10520296409061235
Horobin RW (1983) Staining plastic sections: a review of problems, explanations and possible solutions. J Microsc 131:173–186
Huber JD, Parker F, Odland GF (1968) A basic fuchsin and alkalinized methylene blue rapid stain for epoxy-embedded tissue. Stain Technol 43:83–87. https://doi.org/10.3109/10520296809115048
Humphrey CD, Pittman FE (1974) A simple methylene blue-azure II-basic fuchsin stain for epoxy-embedded tissue sections. Stain Technol 49:9–14. https://doi.org/10.3109/10520297409116929
Jha RK (1976) An improved polychrome staining method for thick epoxy sections. Stain Technol 51:159–162. https://doi.org/10.3109/10520297609116692
Kuznetsova MA, Chaban IA, Sheval EV (2017) Visualization of chromosome condensation in plants with large chromosomes. BMC Plant Biol 17:153. https://doi.org/10.1186/s12870-017-1102-7
Litwin JA (1985) Light microscopic histochemistry on plastic sections. Prog Histochem Cytochem 16:1–84. https://doi.org/10.1016/s0079-6336(85)80001-2
Loussert Fonta C, Humbel BM (2015) Correlative microscopy. Arch Biochem Biophys 581:98–110. https://doi.org/10.1016/j.abb.2015.05.017
Lynn JA (1965) Rapid toluidine blue staining of epon-embedded and mounted ‘adjacent’ sections. Am J Clin Pathol 44:57–58. https://doi.org/10.1093/ajcp/44.1.57
Menovsky T, van den Bergh Weerman M (1999) A modified staining method for connective tissue in semi-thin histological sections of peripheral and cranial nerves. Acta Chir Plast 41:120–122
Morikawa S, Sato A, Ezaki T (2018) A simple, one-step polychromatic staining method for epoxy-embedded semithin tissue sections. Microscopy 67:331–344. https://doi.org/10.1093/jmicro/dfy037
Roberts IM, Hutcheson AM (1975) Handling and staining epoxy resin sections for light microscopy. J Microsc 103:121–126. https://doi.org/10.1111/j.1365-2818.1975.tb04543.x
Sato T, Shamoto M (1973) A simple rapid polychrome stain for epoxy-embedded tissue. Stain Technol 48:223–227. https://doi.org/10.3109/10520297309116628
Trump BF, Smuckler EA, Benditt EP (1961) A method for staining epoxy sections for light microscopy. J Ultrastr Res 5:343–348. https://doi.org/10.1016/s0022-5320(61)80011-7
Twort FW (1924) An improved neutral red light green double stain, for staining animal parasites, micro-organisms and tissues. J State Med 32:351–355
Wells B, La Cour LF (1971) A technique for studying one and the same section of a cell in sequence with the light and electron microscope. J Microsc 93:43–48. https://doi.org/10.1111/j.1365-2818.1971.tb02262.x
Acknowledgements
We are grateful to O. Shirokova for technical support. The work was supported by Russian Science Foundation (Grant 18-14-00195).
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Manskikh, V.N., Sheval, E.V. An adaptation of Twort’s method for polychromatic staining of epoxy-embedded semithin sections. Histochem Cell Biol 153, 121–127 (2020). https://doi.org/10.1007/s00418-019-01836-x
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DOI: https://doi.org/10.1007/s00418-019-01836-x