Large block embedding and "pop-off" technique for immunoelectron microscopy

Area of Science:

• Electron Microscopy
• Immunohistochemistry
• Cell Biology

Background:

• Immunoelectron microscopy (IEM) is crucial for visualizing cellular structures and antigen localization.
• Conventional IEM processing can be challenging for identifying sparse cellular components.
• Improved embedding and sectioning techniques are needed for enhanced IEM analysis.

Purpose of the Study:

• To describe a new IEM technique utilizing large plastic blocks for improved sectioning.
• To evaluate the efficacy of this technique in locating sparse cells and tissue structures.
• To assess the compatibility of different acrylic plastics with this novel method.

Main Methods:

• A large block (6 x 12 mm) embedding technique for IEM was developed.
• Re-embedding of semithin (3-micron) sections was performed directly for thin sectioning.
• Three acrylic plastics (Lowicryl K4M, LR Gold, LR White) were tested for compatibility.

Main Results:

• The large block re-embedding technique facilitated the localization of sparse cells and tissue structures.
• Lowicryl K4M and LR Gold demonstrated excellent ultrastructural morphologic preservation and antigenicity retention.
• These two plastics were successfully adapted to the large block re-embedding technique, comparable to conventional methods.

Conclusions:

• The described large block re-embedding technique is effective for IEM, particularly for sparse targets.
• Lowicryl K4M and LR Gold are suitable embedding media for this technique, ensuring high-quality ultrastructural preservation.
• Technical challenges were noted with LR White, suggesting it is less optimal for this specific application.