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Embedding and sectioning of samples for histological analysis
- 1. Improved Medical Education in Basic Sciences for Better Medical Practicing ImproveMEd
- 2. Embedding and sectioning of samples for histological analysis
- 3. Fixation Embedding Sectioning Staining The main steps in preparation of tissue for histological analysis
- 4. - tissue prepared for microscopic examination Histological slide
- 5. Labeling and transferring the sample
- 6. Dehydration fixation rinsing (water) dehydration alcohol-based fixative Bouin’s fixative For LM 70 % ethanol 96 % ethanol 100 % ethanol sensitive tissues, large samples 50% ethanol 70 % ethanol 96 % ethanol 100 % ethanol postfixation dehydration From 5 % to 100 % ethanol or acetone For EM
- 7. Ethanol Xylene Paraffin Light microscope Propylene oxyde Plastic resins Electron microscope
- 10. Embedding Block casting - molds
- 11. Automated sample processing - from fixation until paraffin infiltration - block forming under human control - sectioning - manually - staining
- 12. Other embedding media Applied if: - The reagents used in paraffin method remove or damage studied tissue components (e.g. lipids) - Heat during paraffin embedding affects the tissue or enzymes - Embedding media isn’t sufficiently hard for the tissue - Thinner sections are needed Synthetic resins Frozen sections
- 13. Embedding samples for transmission electron microscopy Plastic resins - embedding medium solvent is gradually replaced by embedding medium
- 14. Fixation Embedding Sectioning Staining The main steps in preparation of tissue for histological analysis
- 18. Straightening and fixing the sections on the slides
- 21. Serial sections
- 22. Preparation for staining Deparaffination – immersing the slides into xylene (paraffin solvent) – paraffin is removed Rehydraton – immersing the slides in ethanol of descending concentration, and then in water
- 23. Sectioning for electron microscopy (ultramicrotome) Semi-thin sections 0.1 – 2.5 µm - light microscope analysis Ultrathin sections 8 – 100 µm
- 24. Sectioning
- 25. Outer grid diameter = 3 mm Sectioning
- 26. Fixation Embedding Sectioning Staining The main steps in preparation of tissue for histological analysis