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Histopath Lec > tissue processing technique (tissue prepartion and fixation) > Flashcards

tissue processing technique (tissue prepartion and fixation) Flashcards

1
Q

Method of fresh tissue examination

A

Teasing/dissociation
Squash preparation (crushing)
Smear preparation
Frozen section

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2
Q

Selected tissue specimen is immersed in a watch glass containing isotonic solution, carefully dissected and examined under the microscope.

A

Teasing/dissociation

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3
Q

Small piece of tissue < 1mm in diameter is forcibly compressed between two slides or with a cover slip.

A

Squash preparation (crushing)

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4
Q

useful in cytological examination.

A

Smear preparation

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5
Q

Smear preparation

A

Streaking
Spreading
Pull apart
Touch preparation

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6
Q

Specimen is directly and gently applied on a slide in zigzag motion using applicator stick or wire loop.

A

Streaking

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7
Q

Specimen is gently spread in slide into a moderately thick film.

A

Spreading

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8
Q

Suitable specimen for spreading smear preparation

A

Sputum
bronchial aspirates
thick mucoid secretion

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9
Q

A drop of secretion or sediment upon 1 slide and facing it to another slide.

A

Pull-apart

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10
Q

2 slides are pulled apart in a single uninterrupted motion.

A

Pull-apart

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11
Q

Specimen suitable for pull apart smear preparation

A

Serous fluid
sputum
GIT secretion
Blood preparation

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12
Q

Freshly cut tissue is brought into contact and pressed on the surface of a clean glass slide.

A

Touch preparation

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13
Q

Used in rapid tissue examination (cryostat procedure).

A

Frozen section

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14
Q

For rapid diagnosis in cases of emergency.

A

Frozen section

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15
Q

Tissue is hardened by freezing, cut frozen, and stained for microscopic examination.

A

Frozen section

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16
Q

The most critical step in histotechnology

A

Fixation

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17
Q

Defined as the alteration of tissues by stabilizing protein so resist changes such as degeneration, decomposition, putrefaction and tissue distortion.

A

Fixation

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18
Q

brings about crosslinking of proteins which produces denaturation or coagulation of proteins so that the semifluid state is converted into semisolid state.

A

fixative

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19
Q

To preserve the tissue in as life like manner as possible.
Preservation of morphologic and chemical integrity of tissue.

A

Primary aim of fixative

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20
Q

Hardening and solidification.
To protect from trauma or further handling, for easier cutting during gross examination.

A

Secondary aim of fixative

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21
Q

Main factors involved in fixation

A

Hydrogen ion concentration
Temperature
Thickness of section
Osmolality
Concentration
Duration of fixation

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22
Q

Temperature for
Routine
Electron microscopy

A

Room temp
0-4C

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23
Q

Hydrogen ion concentration for fixation

A

pH between 6 and 8

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24
Q

Thickness of section for electron microscopy
light microscopy

A

1-2 mm (squared)
2 cm(squared)

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25
Osmolality for fixation
Slightly hypertonic solution (400-450 mOsm)
26
Concentration of formalin for fixation Routine Immuno-electron microscopy
10% 0.25%
27
Duration of fixation for specimen obtained or remained in fixative over weekend electron microscopy
2-6 hours 3 hours
28
Practical consideration of fixation
Speed Rate of penetration Volume Duration of fixation
29
Rate of penetration of formalin
1mm/hour
30
volume of fixation most effective of tissue fixation
20 x tissue volume
31
Fixation can be hasten by using?
heat vacuum agitation microwave
32
Factors that enhance fixation
Size and thickness - thinner agitation moderate heat (37C)
33
Factors the slows down fixation
Size and thickness - larger Presence of mucus, blood, and fats cold temperature
34
What are the simple chemical fixatives
Aldehyde fixative Metallic fixative Alcohol-based fixative Potassium dichromate fixative Picric acid fixative Glacial acetic acid fixative Acetone fixative Osmium tetra oxide
35
What are the compound chemical fixatives
Micro-anatomical fixative Cytological fixative Histo-chemical fixative
36
What are the aldehyde fixatives
Formaldehyde (formalin) 10 % formol saline 10% neutral buffered formalin Formol-corrosive alcoholic formalin glutaraldehyde
37
is a gas but is soluble in water to the extent of 37-40% w/v.
formaldehyde
38
formaldehyde is a gas but is soluble in water to the extent of 37-40% w/v. this solution of formaldehyde in water is called ___ or _____
formalin full strength formalin
39
one of the commonly used fixative in all laboratories since it is cheap, penetrates rapidly, and does not over harden the tissues.
formalin
40
Time required for fixation at toom temperature for small biopsies at 65C fixation occurs in
12 hours 4-6 hours 2 hours
41
Recommended for CNS tissues and general post-mortem tissues.
10 % formol
42
Ideal for most staining technique including silver impregnation.
10 % formol
43
Best fixative for tissue containing iron pigments.
10 % neutral buffered formalin/phosphate buffered formalin
44
Recommended for post-mortem tissues.
Formol-corrosive (formol-sublimate)
45
Fixes lipids escpecially neutral fats and phospholipids No need for ‘’washing out’’ steps
Formol-corrosive (formol-sublimate)
46
Gendré’s fluid is a?
Alcoholic formalin fixative
47
Used to fix sputum specimen Can be both fixative and dehydrating agent
Alcoholic formalin fixative
48
Suitable for in light microscopy in resin embedding and sectioning for electron microscopy.
Karnovsky’s fixative
49
it is used alone or in combination with osmium tetroxide for electron microscopy.
Glutaraldehyde
50
Recommended for enzyme histochemistry and electron microscopy
Glutaraldehyde
51
What are the metallic fixative
Mercuric chloride Zenkers fluid Zenker-formol Heidenhain's susa B-5 fixative Chromic acid Orth's fluid Lead fixative
52
most common metallic fixative
Mercuric chloride
53
rarely used alone because it causes shrinkage of the tissue. Produces black precipitates (mercuric deposits).
Mercuric chloride
54
Mercuric acids are removed by immersing the tissue before staining to Lugol’s iodine for 5 mins and 5 % Na thiosulfate for 5 mins this process is known as
Dezenkerization.
55
it fixes the cytoplasm without precipitation.
Potassium dichromate
56
Preserves lipids and mitochondria
Potassium dichromate
57
precipitate all proteins excellent for glycogen demonstration suitable for aniline dyes
Picric acid
58
Examples of Picric acid
Bouin's solution Brasil's Icoholic picroformol fixative
59
solidifies at 17˚C it causes the cells to swell hence can never be used alone but should be used with fixatives causing cell shrinkage.
Glacial Acetic acid
60
Alcohol-based fixatives
100 % methyl alcohol 95 % isopropyl alcohol 70 – 100 % ethyl alcohol Carnoy’s fuid
61
used as both fixative and dehydrating agent
Alcohol
62
preserves some proteins in relatively undenatured state so that it can be used for immunofluorescence or some histochemical methods to detect certain enzymes.
ethanol
63
used for fixing blood and bone marrow smears.
methyl alcohol
64
it is a strong oxidizing agent ( pale yellow powder)
Osmium tetroxide
65
used for neurological tissues. e.g. myelin sheath.
Osmium tetroxide
66
Example of Osmium Tetroxide
Flemming's solution Flemming's solution without acetic acid
67
use in cold temperature demonstrates some tissue enzymes like phosphatases and lipases
Acetone
68
fixative agent for rabies diagnosis (Negri bodies)
Acetone
69
such fixatives preserves the anatomy of the tissue.
MICROANATOMICAL FIXATIVES
70
MICROANATOMICAL FIXATIVES
10% formalin heidenhain susa zenkers fluid zenker formal (helly fluid) Bouin's fixative B5 fixative
71
Recommended for fixing small pieces of liver, spleen, connective tissue fibers and nuclei.
Zenker's fluid
72
Suitable for pituitary gland, bone marrow and blood containing organs such s spleen and liver.
Zenker-Formol (Helly’s solution)
73
Recommended for pituitary and embryo biopsies.
Bouin’s fixative
74
Recommended for skin tumor biopsies.
Heidenhain Susa solution
75
Commonly used for bone marrow biopsies.
B5 fixative
76
such fixation are used to preserve intracellular structures or inclusion.
CYTOLOGICAL FIXATIVES
77
Nuclear fixatives
Carnoy's fluid New comer's fluid Flemming's fluid
78
Cytoplasmic Fixatives
Orth's fluid Flemmings fluid withou acetic acid Regard's fluid
79
The most rapid fixative. Recommended for fixing chromosome, lymph glands and urgent biopsies.
Carnoy’s fluid
80
Recommended for fixing mucopolysaccharides and nuclear proteins.
Newcomer’s fluid
81
Serves as both nuclear and histochemical fixatives.
Newcomer’s fluid
82
Recommended for tissue necrosis Preserves myelin sheath better Demonstrate rickettsiae
Orth’s fluid
83
Recommended for demonstration of chromatin, mitochondria, mitotic figures, golgi bodies, RBC and colloid containing tissues.
Regard’s fluid
84
fixative used to preserve the chemical nature of the tissue for further analysis. Used in frozen section
HISTOCHEMICAL FIXATIVES
85
HISTOCHEMICAL FIXATIVES
Absolute ethyl alcohol Acetone 10% formol saline Newcomer's fluid
86
Physical method of fixation
Heat fixation Microwave fixation Freeze drying and freeze substitution
87
Heat primarily used to accelerate other forms of fixation.
Heat fixation
88
What fixation Thermal coagulation of tissue proteins for rapid diagnosis. Usually employed for frozen tissue section and bacteriologic smears.
Heat fixation
89
What fixation increase the movement of molecule to accelerate fixation.
Microwave fixation
90
Useful in preserving neurochemical substances in brain such acetylcholine.
Microwave fixation
91
Microwave fixation treated tissue at what temperature
50C
92
rapid freezing of tissue (-160 to -180 C) and then dehydrated in a vacuum at -30 C to -40 C to remove ice water molecules.
Freeze drying
93
fixed in Rossman’s formula or in 1 % acetone to dehydrate the tissue.
Freeze substitution

Histopath Lec (3 decks)

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