STEPS OF TISSUE PROCESSING in pathology for slide

steps involved in the tissue processing

1. STEPS OF TISSUE PROCESSING DR.R.VENKATESH PATHOLOGY PG 1 ST YEAR MMCHR I

2. AIMS AND PRINCIPLE Aim: The basic aim is to provide sufficient rigidity so it can cut into thin section for microscopic study. Principle: Is to remove the water within the tissue and impregnated with another medium.

3. ESSENTIAL STEPS

4. FACTORS THAT INFLUENCE TISSUE PROCESSING Size of the tissue sample: The smaller size, the better processing Agitation: Facilitate the contact of tissue with fresh solution

5. Heat: Increase the better penetration Viscosity: The higher viscosity, lower the penetration Vacuum: Negative pressure to remove trapped air in the tissue

6. DEHYDRATION Every tissue contains some amount of free or unbound water molecules It is necessary to remove the water from tissue The sharp concentration gradient between the tissue and the dehydration agent may cause rush of fluid, and this may damage

7. So the dehydration should be done gradually from low concentration to the high concentration The tissue should kept in dehydration solution for optimum time Too little time may insufficient to remove the water molecules.

8. DEHYDRARTING AGENTS Ethanol alcohol Methylated spirit Methanol Butyl alcohol Isopropyl alcohol Other than alcohol: dioxane, ethylene glycol and acetone.

9. CLEARING It’s the next step, after removal of water molecules from the tissue The clearing agent should miscible with both dehydrating agent and the embedding medium The refraction index of clearing agent is similar to the tissue

10. SELECTION OF THE APPROPRIATE CLEARING AGENT : Type of tissue Type of processor Processing condition Speed of removal of dehydrating agent Ease of replacement by molten wax Safety factors.

11. CLEARING AGENTS Xylene Toluene Chloroform Esters Cedarwood oil

12. PROPERTIES XYLENE TOLUENE CHLOROFORM ESTERS TISSUE SHRINKAGE Yes Yes Minimum No TISSUE HARDENING Yes No No No INFLAMMABLE Yes Yes No Yes HARMFUL EFFECT Irritant but less harmful Irritant Dangerous toxic gas is liberate Safe COST Cheap Cheap Very expensive High cost

13. INFILTRATION AND EMBEDDING This is the next step after clearing The tissue space is infiltrate embedding medium Usually molten wax is used After cooling in room temperature, the molten wax is solidified to provide support for thin section cutting

14. IMPREGNATING MEDIUM An ideal impregnating medium should have following qualities Miscible with clearing agent Liquid in higher temperature and solid in room temperature Homogenous and stable Non toxic and cheap Transparent Fit for section cutting

15. THE TIME AND NUMBER OF CHANGES DEPENDS ON Size of the tissue Type of the tissue Type of the clearing agent Type of the processing

16. TISSUE PROCESSING METHODS

17. TIME SCHEDULE FOE OVERNIGHT

18. THANK YOU