Preparation of fungi for ultrastructural investigations and immunogoldlabelling

Area of Science:

• Microscopy and Imaging
• Biological Sciences
• Mycology and Plant Pathology

Background:

• Accurate electron microscopic analysis relies on optimal sample preparation.
• Degeneration and artifacts can compromise the integrity of biological samples during observation.
• Specific protocols are needed for preparing complex biological materials like fungi and infected plant tissues.

Purpose of the Study:

• To present comprehensive methods for preparing fungi and plant tissues infected with fungi for transmission electron microscopy (TEM).
• To detail techniques that minimize degeneration and prevent artifacts during sample preparation for TEM.
• To provide protocols for advanced preparation techniques, including cryofixation and immunogold labeling.

Main Methods:

• Chemical fixation techniques for biological samples.
• Cryofixation methods, with a focus on high-pressure freeze fixation.
• Freeze substitution protocols for cryofixed samples.
• Embedding of samples in suitable resins.
• Immunogold labeling of ultrathin sections for enhanced visualization.

Main Results:

• Established protocols for preparing fungal and plant tissues for TEM.
• Demonstrated methods to preserve cellular structures and prevent artifacts.
• Included detailed procedures for high-pressure freeze fixation and subsequent processing.

Conclusions:

• Optimal sample preparation is essential for reliable transmission electron microscopy of fungi and infected plant tissues.
• The presented methods, including cryofixation and immunogold labeling, enable high-resolution imaging of biological samples.
• These protocols contribute to the accurate study of fungal-plant interactions at the ultrastructural level.