Histology

• Source of tissue specimen

  • Biopsy
  • Autopsy (necropsy)
• Biopsy


  • Histological sample taken from living body (i.e through operation) in order to diagnose the disease
  • Should contain both the affected parts and intact parts of tissues
  • Intact parts should have appropriate size for fixation (1 cm)
  • Fixed immediately without washing
  • Made with sharp scalp and avoid any destruction to piece of tissue

  
• Autopsy (necropsy)


  • Histological sample from determined (dead body)
• Tissue preparation/processing steps


  1. Fixation

  2. Dehydration

  3. Clearing

  4. Infiltration

  5. Blocking

  6. Sectioning

  7. Staining
• Fixation


  • Prevent putrefaction of tissue by saprophytic bacteria
  • Prevent autolysis by lysozymes
  • Keep tissue components as possible as it in the living body
• Types of fixatives


  • 10% neutral buffered formalin
  • Ethyl alcohol
  • Carnoys solution
  • Bouins solution
• Dehydration


  Removing the water from tissue by gradual concentration of ethyl alcohol (70%, 80%, 90%, 95%, 100%)
• Clearing


  1. Replacement of alcohol by xylol or (chloroform)

  2. Aims to remove the dehydration agent and make tissue appears clear
• Infiltration


  1. Replace the xylene with paraffin

  2. Harden the tissue
• Blocking


  1. Done with liquid paraffin (56°C)

  2. 2 L shape metal pieces

  3. Stay in room temp. for 15 min

  4. Kept in refrigerator over night
• Sectioning


  1. Slicing the tissue into thin slices 5-6 micrometer (thickness of tissue slices)

  2. Water bath 37°C

  3. Keeping slides in over 37°C over night or 60°C for 2 hr

  
• Staining


  1. Dewaxing removing paraffin from tissue by with xylene

  2. Hydration in descending concentration of ethanol alcohol (100%, 95%, 90%, 100%)

  3. Apply haematoxylin nuclear stain

  4. Complete the nuclear stain by bluing: rinse slides in tap water (weak alkaline sol.) till the nuclei appear blue in color

  5. Apply eosin counter stain

  6. Dehydrate,clear, and mount