Histochemistry Of Oral Tissues Question And Answers

Histochemistry Of Oral Tissues

Describe the stages of tissue processing and write about fixation.
Answer:

Tissue processing:

• It is a long procedure and requires 24 hours.
• Stages:

1. Fixation:

• It is an important step in the preparation of tissue for microscopic examination.
• Goals:
  • Preserves the structure.
  • It terminates any ongoing biochemical reaction.
  • It increases the strength and stability of treated tissue.

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• Purposes:
  • Coagulate proteins.
  • Prevents autolysis of cell
  • Fix cell constituents and tissue that withstand subsequent treatment with various reagents with minimum loss of anatomy.
• Steps:
  • Killing tissue.
  • Preservation of tissue.
  • It is done by fixatives, commonly used – 10% formalin.
• Temperature:
  • Done at room temperature.
• Time:
  • It takes several hours to several days.

2. Dehydration:

• After fixing the tissue, it is washed with running water.
• Water cannot mix with paraffin.
• Hence, the water content is to be removed.
• It is done by passing the specimen by increasing the percentage of alcohol.

3. Clearing:

• Alcohol is not miscible with paraffin.
• Thus, the tissue is first cleared of alcohol by a reagent like xylene.
• Xylene is miscible with both alcohol and paraffin.
  • Time:
    • A small piece of tissue is cleared in 0.5 1 hour.
    • Larger/thicker piece in 2-4 hours.

4. Infiltration:

• When xylene has completely replaced alcohol, the specimen is infiltrated with paraffin at 60°C.

5. Embedding:

• Tissue is embedded in a block of paraffin wax with the proper orientation of the specimen.
• Paraffin wax is used as it has hard consistency.

6. Cutting:

• The paraffin wax block is attached to a wooden cube which is clamped onto the microtome.
• It is adjusted to cut the desired piece of the specimen.

7. Mounting:

• The sections are mounted on microscope slides.

8. Staining:

• Mounted slides are stained to give color to a section.

Procedure:

• Put sections in xylene for 3 minutes
• Transfer to the increasing strength of alcohol for 3 minutes
• Wash in running water for 1 minute
• Put in hemotoxylin for 5 to 7 min.
• Wash in running water for 30 sec.
• Wash excess in 1% acid alcohol for 15 sec.
• Wash in running water for 30 sec.
• Give 2-3 dips of ammonia water solution until tissues attain blue color.
• Wash in running water for 30 sec.
• Counterstain it.
• Wash it
• Dehydrate it.