Tissue Processing

Question 1

Explain in short the steps of tissue processing

Answer 1

1) Fixation - prevents autolysis and putrefaction of the tissue while stabilizing it to maintain its cellular structure as closely to as it was while in vivo
2) dehydration - removes free water and unbound fixative from the tissue in preparation for later impregnation with wax
3) clearing - displaces dehydrating solution with a solution which is miscible with the impregnating medium.
4) infiltration - permeates tissue with a support medium which on solidifcation will be of the same hardness as the embedding medium.
5) embedding - orientation of the tissue sample in a support medium to create a tissue block suitable for sectioning and creation of slides.

Question 2

consequences of over dehydration

Answer 2

rmoval of both bound and free water resulting in
- dry briteel tissue
- shrinkage
- parched earth effect
- abnormal staining

Question 3

consequences of under dehydration

Answer 3

impaires the penetration of the clearing reagents and non receptive to paraffin wax
- soft mushy sections
- greasy and explode or separate rapidly when ribbon placed on water bath

Question 4

most used dehydrating agent

Answer 4

ethanol - ethyl alcohol

Question 5

overexposure of tissue to ethanol

Answer 5

causes excessive shrinkage and hardening of the tissue

Question 6

ideal deatures of dehydrating agent

Answer 6

high affinity for water
penetrates quickly
maintains tissue shape and volume
no brittleness or hardening of tissue
complete dehydration
cheap

Question 7

what can be used instead of ethanol for dehydration

Answer 7

methanol which is harsher but faster acting.
also isopropyl alcohol and acetone. acetone shrinks and hardens tissues more than ethanol.

Question 8

Use of clearing agent

Answer 8

Remove the alcohol used for dehydration as it is not miscible with the impregnation medium. Also has a high refractive indec which renders the tissue transparent.

Question 9

Consequence of prolonged exposure to clearing agent

Answer 9

becomes brittle

Question 10

consequence of inadequate clearing

Answer 10

produces soft mushy tissue

Question 11

types of clearing agents

Answer 11

Aromatic hydrocarbons, aliphatic hydrocarbons and essential oils

Question 12

aromatic hydrocarbons

Answer 12

xyelene or toluene. Most commonly used.

Question 13

aliphatic hydrocarbons

Answer 13

xylene substitutes such as alkanes. They are less agressive than xylene, intolerant for water and incompatabile with some mounting media

Question 14

essnetial oil clearing agents

Answer 14

limonene and cedarwood.

Question 15

What can be used as infiltration medium

Answer 15

• paraffins: made up of petroleum wax, polymers and antioxidants, + others to enhance performance
• carbowax (water soluble)
• celloidin/nitrocellulose (water soluble