Questions and Answers
Which of the following is a chemical fixative?
The purpose of fixation is to prevent autolysis and putrefaction.
True
What is decalcification?
The process of removing calcium salts from hard tissues to make them softer for sectioning.
The primary embedding medium used in routine histology is ____.
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Match the following fixative solutions with their components:
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What are basophilic tissue components?
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Eosin is an example of a basic dye used in staining.
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After fixation, tissues are dehydrated in a series of increasingly concentrated alcohol solutions, ending in _____.
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What is the melting point of paraffin wax?
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Study Notes
Tissue Preparation Steps
- Fixation: Involves placing tissue in chemical solutions to cross-link proteins and preserve structure.
- Dehydration: Transition through increasing concentrations of alcohol to remove water from the tissue.
- Clearing: Alcohol is replaced by organic solvents that mix with both alcohol and paraffin.
- Infiltration: Tissue saturates with melted paraffin to ensure proper embedding.
- Embedding: Paraffin-infiltrated tissue is molded and cooled to form a solid block.
- Trimming: The paraffin block is shaped for sectioning.
Fixatives and Their Combinations
- Common fixatives include formalin, mercuric chloride, acetic acid, picric acid, and glutaraldehyde.
- A mix of fixatives is often used to optimize preservation:
- Bouin’s fluid: Formalin, acetic acid, picric acid.
- Formal sublimate: Formalin, mercuric chloride.
- Helly’s fluid: Formalin, mercuric chloride, potassium dichromate.
- Zenker’s fluid: Acetic acid, mercuric chloride, potassium dichromate.
- Purpose of fixation includes preserving morphology, preventing decay, hardening tissue, solidifying colloidal material, and facilitating staining.
Additional Steps for Hard Tissues
- Decalcification: Hard tissues (e.g., bone, teeth) are treated with agents like nitric acid or EDTA to soften for sectioning.
Dehydration Process
- Tissues are immersed in graded alcohol solutions (50%, 70%, 90%, absolute) for 30-60 minutes each to prepare for embedding in paraffin wax.
Clearing Phase
- Tissues treated with paraffin solvents (xylene/toluene) for 2-3 hours to replace alcohol, rendering tissues translucent.
Embedding Techniques
- Two steps in embedding:
- Impregnation: Tissue is immersed in molten paraffin at 58°–60 °C for 2 hours.
- Casting: Tissue placed in molds with molten paraffin until cooled and solidified.
Microtomy for Section Cutting
- Sections, 5–7 μm thick, are cut using a rotary microtome.
- Sections are affixed to glass microslides and prepared for staining, air-dried or incubated at 37 °C.
Staining Procedure
- Uses basic (e.g., haematoxylin) and acidic (e.g., eosin) dyes to selectively stain tissue components.
- Basophilic components (anionic) appear blue; acidophilic components (cationic) appear pink/orange.
- Combination of haematoxylin and eosin (H&E) is the most common histological staining method.
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Description
This quiz covers the essential techniques for preparing tissues for study in anatomy and physiology. Focus areas include fixation, dehydration, and the importance of chemical solutions in preserving tissue structure. Test your knowledge of tissue preparation methods and their applications in the field.
