[2S] UNIT 5 Dehydration Flashcards by JULIA CASSANDRA RECCION

Q

T/F: Acetone can be used as a Fixative & dehydrating agent

A

T

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Q

Once preserved, the tissue must be processed into a form in which it can be made into thin microscopic sections. This procedure or treatment involves dehydration, clearing, wax infiltration, and embedding.

A

Tissue Processing

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Q

TISSUE PROCESSING

Once preserved, the tissue must be processed into a form in which it can be made into thin microscopic sections. This procedure or treatment involves _______, _________, & _________.

A

dehydration, clearing, wax infiltration, and embedding

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Q

Pre-eminent type of tissue processing treatment
considered to be the most suitable for routine
preparation, sectioning, staining, and subsequent
storage of large tissue samples.

A

Paraffin Wax Method

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Q

It utilizes series of alcohol as dehydrating fluid.

A

Paraffin Wax Method

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Q

is the removal of water from aqueous-fixed tissue

A

Dehyration

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Q

T/F: Most alcohols and paraffin are miscible,
another step, known as clearing, is introduced

A

F; not miscible

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Q

Since most alcohols and paraffin are NOT miscible,
another step, known as _______, is introduced

A

clearing

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Q

CHARACTERISTICS OF AN IDEAL DEHYDRATING SOLUTION

T/F: It should dehydrate rapidly without producing
considerable shrinkage or distortion of tissues.

A

T

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Q

CHARACTERISTICS OF AN IDEAL DEHYDRATING SOLUTION

T/F: It should evaporate very fast.

A

F; should not

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Q

CHARACTERISTICS OF AN IDEAL DEHYDRATING SOLUTION

T/F: It should not dehydrate fatty tissues.

A

F; It should be able to dehydrate even fatty tissues.

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Q

CHARACTERISTICS OF AN IDEAL DEHYDRATING SOLUTION

T/F: It should harden tissues excessively.

A

F; It should not harden tissues excessively.

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Q

CHARACTERISTICS OF AN IDEAL DEHYDRATING SOLUTION

T/F: It should not remove stains.

A

T

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Q

CHARACTERISTICS OF AN IDEAL DEHYDRATING SOLUTION

T/F: It should not be toxic to the body.

A

T

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Q

CHARACTERISTICS OF AN IDEAL DEHYDRATING SOLUTION

T/F: It should not be a fire hazard.

A

T

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Q

Most commonly used dehydrating agent

A

Alcohol

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Q

Commonly Used Dehydrating Agents

A

A ADCTT (adik)

Alcohol (most common)
Acetone
Dioxane
Cellosolve
Triethyl phosphate
Tetrahydrofuran

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Q

It is recommended for routine dehydration of tissues and considered to be the best dehydrating agent.

A

Ethyl Alcohol

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Q

Characteristics: Clear, colorless, flammable fluid

A

Ethyl Alcohol

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Q

Advantages: Fast-acting, mixes with water and many inorganic solvents, penetrates tissue easily, not poisonous and expensive

A

Ethyl Alcohol

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Q

It is primarily used for blood and tissue films, and for smear preparations.

A

Methyl alcohol (methanol)

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Q

It is utilized in plant and animal micro-techniques.

A

Butyl alcohol (butanol)

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Q

Advantage: Slow dehydrating agent producing less shrinkage and hardening than ethanol.

A

Butyl alcohol (butanol)

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Q

Disadvantage: Slow dehydrating agent thus, it is not suitable for rapid tissue processing.

A

Butyl alcohol (butanol)

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Using aqueous fixative in tissue processing

Fixation

What process? 60-70% Ethanol

Dehydration

What process? 2-3 changes of absolute alcohol

Clearing

T/F: The strength of initial alcohol required in each concentration will depend upon the size, and nature of the tissue and fixative used.

T

T/F: Smaller and more delicate tissues require higher concentration and shorter intervals between succeeding ascending grades of alcohol.

F; require lower concentration

Produces shrinkage and make the tissue hard, brittle and difficult to cut. Do not penetrate the deeper parts.

Concentrated Alcohols

What happens if we penetrated the deeper parts of the tissue?

Unequal impregnation & poor sectioning

Length of storage of tissues in alcohol

stored in 70-80% alcohol (not for longer periods of time)

T/F: Prolonged storage in lower concentrations (below 70%) tends to macerate the tissue

T

T/F: Conc. ranging from 70-80% alcohol at very long periods of time does not interfere with the staining properties of the specimen.

F; might interfere

Clear, colorless fluid that mixes with water, ethanol and most organic solvent

Acetone

Advantages • cheap, rapid-acting dehydrating agent which it dehydrate in ½ to 2 hours • more miscible when epoxy resins than alcohol

Acetone

Disadvantages • highly flammable • penetrates tissues poorly • causes brittleness (tissues placed for prolonged period of time)

Acetone

Disadvantages • Most lipids are removed from tissues • extremely volatize and inflammable (limited to small pieces of tissue)

Acetone

• an excellent dehydrating and clearing agent • readily miscible in water, melted paraffin, alcohol and xylol

Dioxane (Diethylene Dioxide)

Advantages • produces less tissue shrinkage • tissues may be placed directly into the solution after washing out

Dioxane (Diethylene Dioxide)

Advantages • Tissues can be left in this reagent for long period of time without affecting the consistency or staining properties of the specimen

Dioxane (Diethylene Dioxide)

Disadvantages • Expensive • extremely dangerous • tissues sections tend to ribbon poorly • Its vapor produce a cumulative and highly toxic

Dioxane (Diethylene Dioxide)

T/F: Dioxane is used routinely

F; SHOULD NOT BRO

T/F: In using dioxane, the laboratory room should be properly ventilated, and all residues should be washed down in the sink.

T

T/F: Dioxane should not be recycled as the risk of creating explosive peroxides increases greatly

T

Time schedule for a dehydration with dioxane: (1st) Pure dioxane solution (2nd) Pure dioxane solution

1 hour

Time schedule for a dehydration with dioxane: (3rd) Pure dioxane solution

2 hours

Time schedule for a dehydration with dioxane: (1st) Paraffin wax (2nd) Paraffin wax (3rd) Paraffin wax

1. 15 mins 2. 45 minutes 3. 2 minutes

T/F: Graupner’s Method should be embedded in mold and cool in water

T

The tissues is wrapped in a gauze bag and suspended in a bottle containing dioxane and a little anhydrous calcium oxide. Water displaced from the tissues by dioxane and in turn absorbed by calcium oxide or quicklime. Dehydration period range from 3-24 hours

Weiseberg’s Method

Dehydration period range of Weiseberg’s Method

3-24 hours

Tissues which have been treated with a chromate fixative, e.g. Regaud’s or Moller’s fluid, should be thoroughly washed in running tap water prior to treatment with dioxane in order to remove the chromate.

Dioxane (Diethylene Dioxide)

CELLOSOLVE • Cellosolve dehydrates rapidly • Stored in it for months without producing hardening or distortion.

Ethylene glycol monoethyl ether

CELLOSOLVE CAUTION: • Combustible at ______ • Toxic

110-120 F

CELLOSOLVE If cannot be avoided ________ ______ ______ _____ should be used instead of ethylene based glycol ethers

propylene based glycol ether

Removes water very readily and produces very little distortion and hardening

Triethyl Phosphate

• Used to dehydrate sections and smears following certain stains • Produce minimum shrinkage

Triethyl Phosphate

• Both dehydrates and clears tissues • Dissolve many substances including fats

Tetrahydrofuran (THF)

• Miscible with lower alcohols, ether, chloroforms, acetone, benzene • May be used for demixing, clearing and dehydrating paraffin section

Tetrahydrofuran (THF)

Most staining procedures give improved results with _________

Tetrahydrofuran (THF)

T/F: THF is toxic, should be in a well-ventilated room & avoided if possible.

T

Many of these dehydrating agents are alcohols of various types that are generally used in increasing strengths to remove aqueous tissue fluids with little disruption to the tissue caused by ________ ______.

diffusion currents

ODD ONE OUT: REAGENT TIME 70% Alcohol Alcohol-I Xylol-I 10% Formalin

10% Formalin kc sha lng 1 hr Paraffin-I & II - 2 hrs d rest 1:30 h

T/F: The dehydrating schedule varies as per the fixative used in the process of Fixation.

T

In case of Carnoy’s fluid and other alcoholic fixatives, the tissues are directly transferred to?

90% or 95% or even Absolute alcohol

What alcohol is used for soft tissues like an embryo and is followed by a graded treatment of the alcohol to prevent tissue shrinkage?

50% Alcohol

Tissues fixed in ____ ______ are transferred to 90% alcohol because lower grades of alcohol may cause swelling of the fibrous tissues.

Susa fluid

Dehydration sequence for specimens not more than 4mm thick

• 70% ethanol 15 min • 90% ethanol 15 min • 100% ethanol 15 min • 100% ethanol 15 min • 100% ethanol 30 min • 100% ethanol 45 min

QUALITY CHECK FOR ABSOLUTE ALCOHOL To ensure that the final bath of the alcohol containing the absolute alcohol or ethanol is free of water, a layer of _______ _____ ________ which should be approx. 2-2-5 cm thick, covered by filter paper to prevent the contamination, is employed in dehydrating vessel.

Anhydrous copper sulfate

QUALITY CHECK FOR ABSOLUTE ALCOHOL Color of alcohol if it gets diluted

Blue

QUALITY CHECK FOR ABSOLUTE ALCOHOL Color of alcohol if it gets diluted

Blue