Laboratory Protocols

• Cryotomy
• Histological Staining Techniques
  • Index
• Histological fixatives
• Peroxidase-anti-peroxidase Technique
• Preparation of tissue section adhesives
• Processing tissues for light microscopy (basics) (PDF 1.1mb)
• Processing tissues for light microscopy (detailed)
• Sample collection guidelines
• Setting up the light microscope
• Specimen preparation for electron microscopy

Histological Staining Techniques

Pearse, 1960.
Do not celloidinise slides. Use a positive control section. Gelatinised slides may be used.

• Dewax sections, rinse in alcohol, rinse in water.
• Filter carbol fuchsin into a coplin jar. Note: If the carbol fuchsin solution has produced a lot of precipitate it should be discarded and a fresh solution prepared.
• Stain at 56^oC for 3 hours.
• Wash well in tap water.
• Differentiate in acid alcohol (1% HCl in 70% alcohol) in a coplin jar until excess background staining is removed, 1 - 3 minutes.
• Wash in tap water.
• Counterstain with 0.25% methylene blue in 1% acetic acid - 1 minute.
• Wash in tap water.
• Rinse in alcohol.
• Clear and mount.

Results:

Lipofuscin - deep magenta red.
Nuclei - blue.
Background - pale pink to pale blue.
Red blood cells - pale pink.

Carbol fuchsin
Dissolve 1g of *basic fuchsin in 10mls of 100% alcohol and add 100mls of 5% aqueous phenol. Mix well for several hours and filter before use.

*Synonyms basic fuchsin / basic red 9 / pararosaniline = C.I. 42500

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