Unit 3.3. Clearing

dealcoholizationproce

Clearing is also know as

Clearing

process wherein alcohol / dehydrating fluid is removed from the tissue and replaced with an intermediate solvent that is fully miscible with both ethanol and paraffin wax

ethanol & paraffin wax

clearing agents must be fully miscible with both _________ and __________

clearing

transition step between dehydration and infiltration

refractive index

clearing agents should have high _____________

at least 10:1

clearing agents volume to tissue ration

1. To make tissue, embryos, and parasites transparent

2. For dealcoholization of tissue preparatory to wax infiltration

3. For dealcoholization of stained sections prior to Permount, Clarite, or Canada balsam

Applications of Clearing

aniline dyes

Clearing agents should not dissolve _____________

1. Xylene

2. Chloroform

3. Benzene

4. Toluene

5. Cedar wood oil

6. Aniline oil

7. Methyl benzoate, Methyl salicylate, & Amyl acetate

8. Clove oil

9. Carbon tetrachloride

10. Other clearing agents

    1. Oil of Bergamont

    2. Oil of Oreganum

    3. Oil of Wintergreen

    4. Carbon disulfide

    5. Carbone xylene

    6. Terpineol

    7. Phenol

    8. High test aviation lead-free gasoline

Enumerate CLEARING AGENTS

xylene

CLEARING AGENTS

• most common & most rapid

• colorless

• miscible with alcohol & paraffin wax

nervous tissue

xylene is not suitable for __________

urgent biopsies ;

15-30 mins

xylene is suitable for ____________, which it clears within _________

milky

xylene becomes _____ when incompletely dehyrated tissues is mixed with it

Chloroform

CLEARING AGENTS

• causes minimum shrinkage and hardening of tissues

• not routinely used

• tough tissues

  • skin

  • fibrinoid

  • decalcified tissues

• nervous tissues

  • brain

• lymph nodes

• embryos

Chloroform is best used for

liver ;

transparent

Chloroform is toxic to ______ upon prolonged exposure and it does not make the tissue ___________

Wrap tissue in gauze

With chloroform, tissues tend to float. what is the remedy?

benzene

CLEARING AGENTS

• rapid acting (5-10 mins), thus used for rapid biopsies

• substitute for xylene

carcinogenic

excessive exposure to benzene may be extremely toxic and may become ___________

aplastic anemia

excessive exposure to benzene may damage the bone marrow resulting in _______________

Toluene

CLEARING AGENTS

• substitute for xylene and benzene for clearing both during embedding and mounting processes (but not as fast as them)

• tend to acidify and damage the pH of the solution

Cedar wood oil

CLEARING AGENTS

• extremely slow clearing agent

• CNS tissues

• cytological studies

• particularly smooth muscles and skin

Cedar wood oil is recommended for

milky;

filtered

Cedar wood oil becomes ________ upon prolonged storage, and should be _________ before use

2-3 days

clearing time of cedar wood oil

Aniline oil

CLEARING AGENTS

• recommended for embryo, insects, and fine, very delicate specimens

• due to its ability to clear 70% alcohol without excessive tissue shrinkage and hardening

Methyl benzoate, Methyl salicylate, & Amyl acetate

CLEARING AGENTS

• very slow acting clearing agent

• used for double embedding technique

clove oil

CLEARING AGENTS

• tissues tend to become adulterated and when used, the wax impregnation is slow and difficult

Carbon tetrachloride

CLEARING AGENTS

• properties and disadvantages are similar to chloroform

• dangerous to inhale due to its toxicity

1. Oil of Bergamont

2. Oil of Oreganum

3. Oil of Wintergreen

4. Carbon disulfide

5. Carbone xylene

6. Terpineol

7. Phenol

8. High test aviation lead-free gasoline

Enumerate OTHER CLEARING AGENTS

Oil of Bergamot & Oil of Oreganum

OTHER CLEARING AGENTS

• for clear smooth muscles to skin

Oil of Wintergreen

OTHER CLEARING AGENTS

• for delicate tissues

Carbon disulfide

OTHER CLEARING AGENTS

• for smooth muscle

• has a foul odor

Carbon xylene

OTHER CLEARING AGENTS

• for fryable tissue

terpineolPge

OTHER CLEARING AGENTS

• for eye specimens

• has a faint odor

• low evaporation rate

phenol

OTHER CLEARING AGENTS

• good substitute for cedar wood oil

High test aviation lead-free gasoline

OTHER CLEARING AGENTS

• excellent clearing agents

15-30 mins

Clearing time of xylene

15-60 mins

Clearing time of benzene

Clearing time of toluene 1-2 hrs