Using Antibodies in Microscopy: A Guide to Immunohistochemistry. Part 3: Post-Embedding Electron Microscopy Techniques.

The article focuses on the use of immunohistochemistry (IHC) with antibodies in post-embedding electron microscopy (EM) techniques to localize proteins at ultrastructural levels. It outlines the critical balance required between tissue fixation, embedding, and antibody staining to preserve cellular ultrastructure while enabling effective antibody binding, recommending specific fixatives like glutaraldehyde and formaldehyde, and IHC-friendly resins such as LR White or Lowicryl. The article discusses labeling strategies using electron-dense markers like colloidal gold for visualization, methods to enhance contrast without compromising antibody binding (e.g., uranyl acetate en bloc staining and osmium tetroxide vapor post-labeling), and considerations for 3D EM imaging. Overall, it emphasizes protocol optimization to achieve precise protein localization in electron microscopy while maintaining structural integrity.