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Fixation Flashcards

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1
Q

What does non-additive fixation means?

A

Fixing agent is not incorporated into the tissue

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2
Q

What does additive fixation means?

A

chemical constituent of the fixative is taken in and becomes part of the tissue

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3
Q

What is the mechanism of additive fixation?

A

forming cross-links or molecular complexes and giving stability to the protein

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4
Q

What is the mechanism of non-additive fixation?

A

Alters the tissue composition and stabilizes the tissue by removing the bound water attached to H-bonds of certain groups within the protein molecule

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5
Q

Fixation temperature for Electron Microscopy and Histochemistry

A

4°C

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6
Q

Fixation temperature using autotechnicon

A

40°C

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7
Q

Temperature for manual fixation

A

Room temperature

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8
Q

Penetration rate of formalin

A

1 mm/hr

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9
Q

Fixative volume for maximum effective fixation

A

20x the volume of the specimen

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10
Q

Enumerate the 5 NUCLEAR fixatives:

A

Carnoy’s
Bouin’s
Newcomer’s
Heidenhain’s Susa
Flemmings with HAc (Acetic acid)

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11
Q

Enumerate the 4 CYTOPLASMIC fixatives:

A

Regaud’s/Moller’s
Orth’s
Formalin with post chroming
Flemmings without HAc (Acetic acid)

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12
Q

How to make 1 Liter of 10% formalin?

A

100 ml of 37%-40% formalin + 900 ml distilled water

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13
Q

Disadvantage of formalin

A

Prolonged storing causes precipitation of formaldehyde to paraformaldehyde

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14
Q

Remedy to prevent the precipitation of formaldehyde

A

Add 10% methanol

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15
Q

Primary goal of fixation

A

Preserve tissue

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16
Q

Secondary goal of fixation

A

Hardens tissue
Protects tissue from trauma of further handling

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17
Q

Why do we need to achieve hardening of tissue?

A

To facilitate easy cutting of specimen into thin slices

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18
Q

Factors involved in fixation: (7)

A
  1. Correct size and thickness
  2. pH
  3. Osmolality
  4. Concentration of fixative
  5. Observe correct volume
  6. Time & duration
  7. Temperature
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19
Q

Factors to be considered when choosing the appropriate fixative: (5)

A
  1. Urgency of the case
  2. Type of tissue to be processed
  3. Tissue structure to be studied
  4. Staining technique
  5. Type of section to be studied
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20
Q

Factors that retard/slow fixation process: (4)

A
  1. Cold temperature
  2. Presence of blood and mucus
  3. Size and thickness of specimen
  4. Presence of Fats
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21
Q

Remedy for presence of blood and/or mucus in the tissue

A

Wash the tissue with NSS

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22
Q

Remedy for specimens with fats

A

Cut the specimen thinly to ensure rapid entry or penetration of fixative

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23
Q

Factors that accelerates fixation:

A
  1. Size and thickness
  2. Heat application
  3. Agitation
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24
Q

Problems encountered during fixation

A
  1. Loss of substances soluble in fixing agents
  2. Inadequate fixation
  3. Over fixation
  4. Presence of Artifacts
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25
Ideal pH of fixative
pH 6-8
26
Ideal osmolality of fixative (by the book)
Slightly hypertonic
27
Ideal osmolality of fixative in real setting
Isotonic
28
Correct size and thickness of specimen
3x2 cm, 3-5mm thick
29
Recommended size for large specimens like lungs
1-2cm
30
Ratio of fixative volume to specimen
15-20:1
31
Which fixative must be 5-10x the volume of the specimen if used a. Mercuric Chloride b. Bouin's c. Formol Zenkers/Zenker's formol d. Osmium tetroxide
D. Osmium tetroxide
32
Formaldehyde/Formalin Fixatives (4)
1. 10% formol saline 2. 10% Neutral Buffered formalin 3. Formol corrosive/Formol sublimate 4. Alcoholic Formalin/Gendres
33
Classification of fixatives according to action/purpose that preserves cell parts
Cytological fixative
34
What are the 2 cytological fixative?
1. Nuclear Fixatives 2. Cytoplasmic fixatives
35
Classification of fixatives according to action/purpose that preserves tissues and cells in general
Microanatomical
36
Classification of fixatives according to action/purpose that preserves chemicals
Histochemical
37
Aldehyde fixatives under Formaldehyde/Formalin: (5)
1. 10% Formalin 2. 10% Formol Saline 3. 10% Neutral Buffered Formalin 4. Formol Corrosive/Formol Sublimate 5. Alcoholic Formalin/Gendres
38
Formaldehyde fixative that is routinely used
10% formalin
39
Formaldehyde fixative that is used for CNS and post mortem tissues. NSS is used as diluent
10% formol saline
40
Formaldehyde fixative that is used for tissues with iron pigments.
10% Neutral Buffered Formalin
41
Composition of 10% Neutral Buffered Formalin
10% formalin + Sodium phosphate (buffer)
42
Formaldehyde fixative that is best for lipids and neutral phospholipids
Formol corrosive/Formol sublimate
43
Composition of formol sublimate
Formaldehyde + Mercuric chloride
44
Formaldehyde fixative that is used for microincineration and sputum specimens
Alcoholic formalin/Gendres
45
Composition of Gendres fixative
Formaldehyde + 95% Ethanol + Glacial HAc +Picric acid
46
Aldehyde fixative that is used for Electron microscopy
Glutaraldehyde
47
Amount of glutaraldehyde used for small tissue fragments/needle biopsies
2.5% Glutaraldehyde
48
Amount of glutaraldehyde used for larger tissues less than 4mm thick
4%
49
Aldehyde fixative that is the smallest aldehyde
Glyoxal
50
Aldehyde fixatives: (3)
1. Formaldehyde/Formalin 2. Glutaraldehyde 3. Glyoxal
51
Metallic Fixatives: (3)
1. Mercuric Chloride Fixatives 2. Chromate Fixatives 3. Lead fixatives
52
Most common metallic fixative
Mercuric Chloride
53
Metallic fixative that is excellent for Trichrome staining & tissue photography
Mercuric Chloride
54
How do you remove excess mercuric fixatives:
Wash using alcoholic iodine
55
Disadvantage of mercuric chloride fixative
Toxic and leaves excess black mercury deposit
56
Mercuric chloride fixative that is used for liver, spleen, connective tissues and nuclei
Zenker's Fluid
57
Mercuric chloride fixative that is used for bone marrow
B5
58
Mercuric chloride fixative that is used for tumor skin biopsies
Heidenhain's Susa
59
Mercuric chloride fixative that is used for pituitary gland, bone marrow, spleen, and liver
Helly's Fluid/Zenker Formol
60
Substitute for mercuric chloride but is also toxic
Zinc sulfate
61
Mercuric Chloride fixatives: (5)
1. Zenker's fluid 2. B5 3. Heidenhain's susa 4. Helly's fluid/Zenker formol 5. Zinc sulfate
62
Chromate fixatives: (4)
1. 1-2% Chromic acid 2. 3% Potassium Dichromate 3. Regaud's Fluid/Moeller's 4. Orth's fluid
63
Chromate fixative that is used for carbohydrate
1-2% Chromic acid
64
Chromate fixative that is used for lipids and mitochondria
3% Potassium Dichromate
65
Chromate fixative that is used for mitochondria, RBC, colloid containing tissues
Regaud's Fluid/Moeller's
66
Chromate fixative that is used for Ricketssiae & other bacteria, tissue necrosis
Orth's fluid
67
Metallic fixative that is used for Acid mucopolysaccharides and tissue mucin
Lead fixative
68
Fixative that is excellent for glycogen demonstration
Picric acid fixative
69
Three (3) uses of Picric acid
1. Fixative 2. Decalcifying agent 3. Stain
70
Fixative that imparts yellow color
Picric acid
71
This is used to wash out excess amount of picric acid fixative
50-70% alcohol
72
Picric acid fixative: (3)
1. Bouin's fluid 2. Brasil's 3. Hollande's
73
Picric acid fixative that is used for for embryos, pituitary biopsies, and endometrial curetting
Bouin's fluid
74
Picric acid fixative that is not recommended for kidney structures and not compatible with Feulgen's stain
Bouin's fluid
75
Picric acid fixative that is excellent for glycogen
Brasil's
76
Picric acid fixative that is used for gastrointestinal tract biopsies and endocrine tissues
Hollande's
77
Alcohol fixatives: (3)
1. 95% Isopropyl Alcohol 2. Carnoy's 3. Newcomer's
78
Alcohol fixative that is used for touch preparations
95% isopropyl alcohol
79
Alcohol fixative that is the most rapid ftissue ixative
Carnoy's
80
Alcohol fixative that is used for chromosomes, lymph glands, and brain tissues
Carnoy's
81
Alcohol fixative that aids in rabies diagnosis together with acetone
Carnoy's
82
Alcohol fixative that is used for mucopolysaccharides and nuclear protein
Newcomer's
83
Uses of Acetone: (2)
1. Fixative 2. Dehydrating Agent
84
Fixative that is used for phosphatases, lipases, and brain tissues
Acetone
85
Fixative that must be used at ice cold temperature
Acetone
86
Fixative that easily evaporates and dissolves fat
Acetone
87
Fixative that is used for electron microscopy and used for myelin and peripheral nerves
1. Osmium Tetroxide 2. Glutaraldehyde 3. Paraformaldehyde

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