4. Taking Samples:
Edge of lesions.
Wall of cysts.
Whole specimen if
small.
Direction, mark
5. Inking the Margins
To mark surgical
margin.
Spread of lesion
Malignancy
Adequacy of
removal
Different colors to
identify margins
6. Fixation:
Specimen bits are
placed in porous
cassettes
Not more than 5mm
thick
In 10% formalin
1mm/hour fixation
~ 6 hour
7. Types of fixative:
Acetic acid
Formaldehyde
Ethanol
Glutaraldehyde
Methanol and Picric acid.
8. Dehydration
A. Definition: removal of water
B. Rationale: for paraffin embedding/sectioning
C. Steps
1. wash out fixative
2. graded series of alcohol
70%, 95%, 100%, 100%
3. replace water by diffusion
4. not too long, not too short
10. Clearing:
• replacing the dehydrating fluid with a fluid that is totally
miscible with both the dehydrating fluid and the embedding
medium.
Choice of a clearing agent depends upon the
following
- Intended processing conditions such as temperature,
vacuum and pressure.
- Safety factors.
- Cost and convenience.
- Speedy removal of dehydrating agent .
- Ease of removal by molten paraffin wax .
- Minimal tissue damage .
11. is the process by which tissues are
surrounded by a medium such as agar,
gelatin, or wax which when solidified will
provide sufficient external support during
sectioning.
Embedding
12. Embedding
• Procedure;
• 1. Place tissue cassette
in melted paraffin
• 2. Fill mold with
paraffin
• 3. Place tissue in
mold
• 4. Allow to cool
14. Precaution while embedding in wax
The wax is clear of clearing agent.
No dust particles must be present.
Immediately after tissue embedding, the wax must be rapidly
cooled to reduce the wax crystal size.
15. There are four main mould systems and associated
embedding protocols presently in use :
• 1- traditional methods using paper boats
• 2- Leuckart or Dimmock embedding irons or metal containers
• 3- the Peel-a-way system using disposable plastic moulds and
• 4- systems using embedding rings or cassette-bases
16. Embedding:
Paraffin block with
embedded tissue
consistency to cut
Paraffin blocks taken
for sectioning