1. Fixation:
* This is the first and most critical step.
* The goal is to preserve the tissue structure and prevent decomposition.
* Common fixatives include formalin, glutaraldehyde, and alcohol.
2. Dehydration:
* Water is removed from the tissue using a graded series of alcohol solutions (usually 70%, 90%, 95%, and 100%).
* This prepares the tissue for embedding in paraffin wax.
3. Clearing:
* Alcohol is removed from the tissue and replaced with a solvent that is miscible with both alcohol and paraffin wax (e.g., xylene, toluene, or Histoclear).
4. Embedding:
* The tissue is infiltrated with melted paraffin wax under vacuum.
* This process allows the wax to permeate the tissue and solidify, creating a firm block that can be sectioned.
5. Sectioning:
* The paraffin block is trimmed and sliced into thin sections (typically 4-10 micrometers thick) using a microtome.
6. Mounting:
* The sections are carefully mounted onto glass slides, usually with a water-soluble adhesive.
7. Deparaffinization:
* The paraffin wax is removed from the tissue sections using a series of solvents (e.g., xylene, toluene).
8. Rehydration:
* The tissue sections are rehydrated through a graded series of alcohol solutions (e.g., 100%, 95%, 90%, 70%, and water).
9. Staining:
* This is where the tissue components are visualized.
* There are numerous stains, each with its own properties and applications.
* Examples include hematoxylin and eosin (H&E) for general morphology, and special stains for specific cell types or structures.
10. Mounting:
* The stained sections are mounted with a coverslip and a mounting medium (e.g., DPX, Permount).
11. Labeling:
* The slides are labeled with relevant information (patient name, date, tissue type, stain).
Note: This is a standard protocol. There may be variations depending on the specific tissue type, staining techniques, and research applications.
