Post-embedding Mammalian Tissue for Immunoelectron Microscopy: A Standardized Procedure Based on Heat-Induced Antigen Retrieval.

Abstract

We describe a standardized method of fixation, antigen retrieval, and image contrasting for post-embedding immunoelectron microscopy. Tissues are fixed with formaldehyde solutions containing Ca(2+) and Mg(2+) ions at pH 7.4 and then at pH 8.5. After dehydration with dimethylformamide, the specimens are embedded in LR-White resin. For antigen retrieval, ultrathin sections are heated in 0.5 M Tris-HCl, pH 9.0, for 1-2 h at 95 °C. After immunogold labeling, the sections are treated with a mixture of tannic acid and glutaraldehyde, with OsO4 solution, and then double-stained with uranyl acetate and lead citrate. The standardized method yields strong and reproducible immunoreactions for many antigens showing excellent image contrast without destruction of fine structures.

DOI: 10.1007/978-1-4939-6352-2_18

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Cite this paper

@article{Yamashita2016PostembeddingMT, title={Post-embedding Mammalian Tissue for Immunoelectron Microscopy: A Standardized Procedure Based on Heat-Induced Antigen Retrieval.}, author={Shuji Yamashita}, journal={Methods in molecular biology}, year={2016}, volume={1474}, pages={279-90} }