Stem Sectioning and Staining Techniques

Stem Sectioning and Staining

Sectioning Techniques:

• Hand Sectioning: Cutting stem sections using a razor blade or sharp knife, typically used for thick stems or when a microtome is not available.
• Microtome Sectioning: Using a microtome to cut thin sections (5-20 μm) of stem tissue, providing higher resolution and more precise control.
• Cryosectioning: Freezing stem tissue and cutting sections using a cryostat, often used for soft or delicate tissues.

Staining Techniques:

• Temporary Stains: Used for quick observation, such as toluidine blue or methylene blue, which can be easily removed.
• Permanent Stains: Used for long-term preservation and observation, such as safranin and fast green, which are more resistant to fading.

Common Stains for Stem Sectioning:

• Safranin and Fast Green: A combination stain for general histology, differentiating between lignified and non-lignified tissues.
• Toluidine Blue: A metachromatic stain for revealing cellulose, pectin, and lignin in plant cell walls.
• Methylene Blue: A basic dye for staining plant cell walls and nucleic acids.

Preparation and Fixation:

• Fixation: Using chemicals like formaldehyde, ethanol, or acetic acid to preserve stem tissue and prevent degradation.
• Dehydration: Gradually replacing water in the tissue with solvents like ethanol or acetone to prepare for embedding.
• Embedding: Infiltrating the tissue with a resin or wax to provide support and stability for sectioning.

Mounting and Observation:

• Mounting Media: Using a medium like Canada balsam or Permount to attach the section to a microscope slide.
• Microscopy: Observing the stained sections using a light microscope to examine stem tissue structure and organization.

Stem Sectioning and Staining

Sectioning Techniques

• Hand sectioning uses a razor blade or sharp knife to cut stem sections, ideal for thick stems or when a microtome is unavailable.
• Microtome sectioning cuts thin sections (5-20 μm) of stem tissue, providing higher resolution and precise control.
• Cryosectioning freezes stem tissue and cuts sections using a cryostat, suitable for soft or delicate tissues.

Staining Techniques

• Temporary stains like toluidine blue or methylene blue are used for quick observation and can be easily removed.
• Permanent stains like safranin and fast green are used for long-term preservation and observation, resisting fading.

Common Stains for Stem Sectioning

• Safranin and fast green combination stain differentiates between lignified and non-lignified tissues in general histology.
• Toluidine blue is a metachromatic stain revealing cellulose, pectin, and lignin in plant cell walls.
• Methylene blue is a basic dye for staining plant cell walls and nucleic acids.

Preparation and Fixation

• Fixation uses chemicals like formaldehyde, ethanol, or acetic acid to preserve stem tissue and prevent degradation.
• Dehydration gradually replaces water in the tissue with solvents like ethanol or acetone to prepare for embedding.
• Embedding infiltrates the tissue with a resin or wax to provide support and stability for sectioning.

Mounting and Observation

• Mounting media like Canada balsam or Permount attach the section to a microscope slide.
• Microscopy uses a light microscope to examine stained sections and observe stem tissue structure and organization.