Fixation and embedding

Area of Science:

• Electron Microscopy
• Immunocytochemistry
• Cell Biology

Background:

• Fixation in electron microscopy presents a challenge, balancing morphological preservation with antigenicity.
• The choice of fixative and embedding resin is critical for immunogold labeling success.
• Labeling can occur pre- or post-embedding, each with specific fixation requirements.

Purpose of the Study:

• To outline optimal fixation and embedding strategies for electron microscopic immunocytochemistry.
• To address the compromise between morphology and antigenicity in electron microscopy.
• To guide the selection of methods for pre- and post-embedding immunogold labeling.

Main Methods:

• Discusses conventional and microwave fixation techniques.
• Explores various embedding resins and their impact on immunostaining.
• Details strategies for pre-embedding immunolabeling with optional refixation.
• Covers post-embedding immunolabeling, emphasizing initial fixative composition.

Main Results:

• Pre-embedding staining allows for mild fixation or unfixed labeling followed by stronger refixation.
• Post-embedding staining necessitates a single fixative that preserves both morphology and antigenicity.
• Embedding resins can influence the outcome of post-embedding immunogold staining.

Conclusions:

• Effective electron microscopic immunocytochemistry relies on carefully chosen fixation and embedding protocols.
• Method selection depends on whether labeling precedes or follows sample embedding.
• Preserving antigenicity alongside morphological detail is key for reliable immunolabeling results.