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Histotechnology > Fixation > Flashcards

Fixation Flashcards

1
Q

What are the functions of a fixative

A

prevent autolysis, stabilize tissue morphology and enhance staining

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2
Q

What are the two major ways that fixatives alter tissue proteins

A

denaturation and formation of crosslinks

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3
Q

What are the four methods of denaturation

A

heat, alcohols, acids and heavy metals

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4
Q

What is the most common kind of cross-linking fixatives

A

aldehydes

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5
Q

What do aldehydes bind to

A

amino groups as well as lysine, cysteine, serine and threonine residues

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6
Q

How does fixation using oxidizing agents work

A

poorly understood but it is known that tissues are stabilized by cross-linking reactive groups in close prozimity

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7
Q

What are coagulant fixatives

A

cause distortion and destruction of fragile cytoplasmic constituents but support proteins. They work quickly

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8
Q

What are examples of coagulant fixatives

A

alcohols, acetone, acid fixatives and metal salts

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9
Q

What are non-coagulant fixatives

A

stabilize tissue morphology by forming bridges between adjacent reactive areas within proteins. Slower

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10
Q

What are examples of non-coagulant fixatives

A

aldehydes and oxidizing reagents

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11
Q

What is an additive

A

chemically binds with the proteins or other tissue component

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12
Q

What is a non-additive

A

disrupting tissue structure without becoming chemically incorporated

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13
Q

What are the main factors that affect fixation

A

temperature, size/thickness/tissue type, time, volume of fixative

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14
Q

What is the minimum fixation time

A

8 hours

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15
Q

What is 100% formalin

A

47-40% formaldehyde in water

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16
Q

What is neutral buffered formalin

A

10% formalin in water

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17
Q

How much formaldehyde is in neutral buffered formalin

A

3.7-4%

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18
Q

What is paraformaldehyde

A

a white powder that precipitates out of concentrated formaldehyde solutions

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19
Q

What type of fixative is formalin

A

additive, non-coagulant, cross-linking

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20
Q

What are the pros and cons of formalin

A

less shrinkage but hardens tissue

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21
Q

What is a limit of glutaraldehyde

A

tissue must be very thin

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22
Q

What type of fixative is glutaraldehyde

A

additive, non-coagulant fixative

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23
Q

When is glutaraldehyde used

A

for electron microscopy

24
Q

Why is acetic acid added to fixatives

A

it preserves nucleoproteins and precipitates DNA, it can lyse RBCs and it swells tissues

25
Q

What is picric acid

A

both a fixative and a stain

26
Q

What type of fixative is picric acid

A

additive coagulant

27
Q

Why must picric acid be avoided if DNA/RNA analysis is anticipated

A

it hydrolyzes nucleic acids

28
Q

What type of dyes does picric acid enhance

A

anionic dyes

29
Q

What must occur after fixation with picric acid

A

the picric acid must be neutralized

30
Q

What is ethanol used for in the histo lab

A

mostly as dehydrating agents

31
Q

What does fixation with alcohols do to tissues

A

leaves tissue hard and shrunken

32
Q

What is ethanol notable for

A

preserving glycogen and urate crystals

33
Q

What type of fixative is osmium tetroxide

A

additive, non-coagulant

34
Q

What is osmium tetroxide used for

A

electron microscopy

35
Q

What makes osmium tetroxide notable

A

it can fix lipids causing them to appear black

36
Q

What fixatives are included in bouins solution

A

formaldehyde, acetic acid and aqueous picric acid

37
Q

What makes bouins solution ideal

A

good for trichromes as the picric acid enhances staining, the acetic acid helps counteract shrinkage, and picric acid leaves the block soft and easy to section

38
Q

Why is it important to wash tissue after fixation with bouins solution

A

the residual picric acid will digest the section over time

39
Q

What are the components of B5

A

mercuric chloride, sodium acetate, formalin and water

40
Q

What is B5 most often used

A

hematopeitic and luphoreticular tissues

41
Q

Why must tissues be stored in 70% alcohol after fixation with B5

A

it is not tolerant

42
Q

What is B-plus

A

a modern version of B5 that uses zinc chloride instead of mercuric chloride

43
Q

What are the components of Clarke’s fluid

A

ethanol and glacial acetic acid

44
Q

What does Clarke’s fluid do

A

preserves nucleic acids, extracts lipids and maintains microanatomical structures

45
Q

What is the storage solution after fixation with Clarke’s fluid

A

95% ethanol

46
Q

What are the components of modified formalin

A

formaldehyde and alcohol

47
Q

What is the benefit of modified formalin

A

it speeds up tissue fixation and begins dehydration

48
Q

What does adding zinc sulfate to NBF do

A

preserves tissue antigenicity with prolonged storage and improves nuclear detail

49
Q

What is proprietary formalin-free fixative

A

usually contain an alcohol and other additives, safer solution than formalin

50
Q

What is melanin

A

a dark brown, granular pigment often demonstrated using the fontana-masson stain and may be removed using potassium permanganate

51
Q

What is formalin pigment

A

amorphous birefringent dark brown or black pigment formed from hemoglobin reacting with formaldehyde in acidic conditions

52
Q

How to remove formalin pigment from a section

A

saturated solution of picric acid and alcohol followed by a running water wash

53
Q

What is mercury pigment

A

a crystalline birefringent brown pigment

54
Q

How to correct mercury pigment

A

use a iodine treatment followed by a sodium thiosulphate treatment

55
Q

What is picric acid pigment

A

stains tissue components an intense yellow colour, rinising the tissue helps remove it

56
Q

What are two examples of exogenous pigments

A

carbon and tattoo ink

Histotechnology (10 decks)

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