PRINCIPLE OF TISSUE PROCESSING

Surgical Specimen

Clinical Details Adequate specimen Proper Fixative

Gross Examination

Description: Specimen weight &

measurement (approx)

Consistency Cut section

Taking Samples:

Edge of lesions. Wall of cysts. Whole specimen if

small. Direction, mark

Inking the Margins

To mark surgical margin.

Spread of lesion Malignancy Adequacy of

removal Different colors to

identify margins

Fixation:

Specimen bits are placed in porous cassettes

Not more than 5mm thick

In 10% formalin 1mm/hour fixation ~ 6 hour

Types of fixative: Acetic acid Formaldehyde Ethanol Glutaraldehyde Methanol and Picric acid.

DehydrationA. 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

DehydrationProcedure; 1. automatic tissue

processor a. overnight 2. Baths: water,

70,95,100,100 % alcohol

3. Clearing agent: 2 baths of xylene

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 .

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

Embedding• Procedure;• 1. Place tissue cassette

in melted paraffin• 2. Fill mold with

paraffin• 3. Place tissue in

mold• 4. Allow to cool

EmbeddingA. Orient tissue

1. cross section

2. longitudinal section

B. Dissection orientation

C. Avoid bubbles

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.

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

Embedding:

Paraffin block with embedded tissue

consistency to cut Paraffin blocks taken

for sectioning