Clearing (Gregorios)

Flashcards reviewing the clearing process in histology, including agents, characteristics, and substitutes for xylene.

What is clearing in histology?

The process of removing alcohol or a dehydrating agent from tissue and replacing it with a substance that dissolves wax.

What are the main purposes of a clearing agent?

To dissolve wax, be miscible with mounting media, and impart optical clarity to tissue.

What is the most commonly used clearing agent for removing fat from tissue?

Xylene

What is/are the clearing agents used when tissue is to be cleared directly from water?

Glycerin and gum syrup

What are the important characteristics of a good clearing agent?

Miscibility with alcohol and paraffin wax, minimal tissue damage, and transparency

What factors influence the choice of a clearing agent?

Type of tissue, processor system, and safety factors

What is the effect of prolonged exposure to most clearing agents?

They cause the tissue to become brittle and difficult to cut.

What is Xylene?

Colorless clearing agent commonly used in histology laboratories

What are some toxicities associated with xylene metabolism?

Central nervous system disorders and respiratory depression.

What are the advantages of using xylene as a clearing agent?

Rapid clearing, miscible with alcohol and paraffin, and does not extract aniline dyes.

What are the disadvantages of using xylene?

It makes tissues excessively hard and brittle if used longer than 3 hours.

Where should xylene be stored?

Flammable liquid storage cabinet

Which clearing agent preserves tissue structure better than xylene and is more tolerant of water?

Toluene

What are the disadvantages of using toluene?

Toxic and more expensive than xylene.

What is an advantage of using Toluene as a clearing agent?

It does not cause tissues to become excessively hard and brittle

Which agent was once a popular clearing agent but is now discouraged due to its carcinogenic properties?

Benzene

What are the advantages of using benzene as a clearing agent?

Rapid clearing and minimal shrinkage

What are the health hazards associated with benzene?

Carcinogenic and may damage bone marrow

Which clearing agent causes less brittleness than xylene but does not make tissues translucent?

Chloroform

What are the advantages of using chloroform?

Minimum shrinkage and hardening

What is a disadvantage of using chloroform?

Relatively toxic to the liver

Which clearing agent is recommended for central nervous system tissues?

Cedarwood oil

What are the advantages of using cedarwood oil?

Very penetrating and minimal shrinkage of tissues.

What are the disadvantages of using cedarwood oil?

Extremely slow clearing agent which is not used for routine usecases

Which agent is recommended for clearing embryos and delicate specimens without excessive shrinkage?

Aniline oil

Which clearing agent causes minimum shrinkage but is not guaranteed in quality and causes tissues to become brittle?

Clove oil

Which clearing agent can be used for embedding tissues but produces considerable tissue hardening and is dangerous to inhale?

Carbon tetrachloride

Which agent can perform dehydration and clearing at the same time, but has an offensive odor?

Tetrahydrofuran

Which clearing agent is miscible with both water and paraffin and allows for embedding within 4 hours after fixation?

Dioxane

Name some xylene substitutes.

Limonene, mineral oil mixtures, vegetable oils and coconut oil

What hampers the use of xylene substitutes?

They are not avilable in commercial quantitites

Where are terpenes found?

Essential oils derived from plants

Name some terpenes.

Turpentine, cedarwood, clove, and lemon oils

What is the effect of terpenes?

They are effective solvents but considered toxic.

Name one of the recommended xylene substitutes from the terpene family.

Limonene

How is limonene obtained industrially?

Steam distillation of orange peel

Why is limonene not universally successful?

Mounting medium does not mix well with limonene

What are the benefits of using orange oil based clearing agents?

Offer clearing action with the lowest hazard rating.

What happens when the orange oils are neither pure nor stable?

They can break down to produce compounds which will interfere with staining procedures.

What health risks are posed by Chlorinated hydrocarbons?

Toxic Chemicals with serious health risks

Which substance is an efficient, non-hazardous, and less expensive substitute for xylene that causes less tissue shrinkage?

Coconut oil

What is one drawback associated with coconut oil?

It tends to solidify at lower temperatures.

How can the issue of coconut oil solidifying at a lower temperature be overcome?

Conduct the clearing procedure in an incubator.

The substitution of which agent gives good tissues, sections and histological slides and is also nontoxic, nonhazardous, nonflammable, bio-degradable, economic, easy to handle, and readily available?

Bleached palm oil

Which clearing agent is likely to acidify in a partially filled vessel?

Toluene

What is a necessary special handling procedure for xylene?

Wash hands thoroughly after handling xylene

Where should xylene be stored?

Store xylene in a flammable liquid storage cabinet

What happens when an incompletely dehydrated tissue is immersed in xylene?

Xylene will become milky

What must be done in order to prevent xylene from subliming and entering the atmosphere?

Keep container tightly closed

Which clearing agent causes a greater shrinkage than xylene does?

Dioxane