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Histopathology Lecture > Mod 5: HISTOPATHOLOGIC TECHNIQUES > Flashcards

Mod 5: HISTOPATHOLOGIC TECHNIQUES Flashcards

1
Q

This is prepared by cutting a thin slice from a small piece of fixed tissue

A

Section

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2
Q

What stain is used for viral inclusions?

A

H&E Staining

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3
Q

What stain is used for Negri Bodies (Rabies)?

A

Seller’s stain

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4
Q

A machine that fixes, dehydrates, clears, and infiltrates the tissue

A

Automatic processors

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5
Q

How many steps are in histopathologic techniques?

A

15 steps

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6
Q

These are samples for the purpose of cytological studies (FNA)?

A

Fine Needle Aspirations

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6
Q

Primary technique used for diagnosis of “Skin Specimens”

A

Punch biopsy

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7
Q

This biopsy is you took a portion of the cell and the surrounding tissue.

Used for bone marrow

A

Core Needle Biopsy

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8
Q

Histological Preparation made from blood, bone marrow, or any fluid such as pleural or ascitic fluid

A

Smears

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8
Q

This involves swabbing, brushing, lavage, washing, scraping, collection of secretions, shavings, and curettings (layman term of raspa)

A

Foliative cytology

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8
Q

Usually applied for small samples such as samples that are easily macerated

A

Whole mount

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9
Q

Whole Mount specimen thickness

A

No more than 0.2-0.5 mm in thickness

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9
Q

Majority of the preparation in histopathology. It is the cutting of the tissue

A

Sections

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10
Q

Sections specimen thickness

A

3-5 mm thick pieces

5 microns thick sections are cut on a microtome

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11
Q

Blocks of tissues taken for processing should be left in __ formalin at __C till processing

This would be fixed in __ hours

A

10% formalin

60C

2 hours

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12
Q

What is the size of the specimen piece to achieve better penetration of fixative

A

1cm

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12
Q

Tissues should be fixed in less than __ hour to avoid biochemical changes

A

less than “1 hour”

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12
Q

These are fixatives that are made up of only one component substance

A

Simple Fixatives

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13
Q

In Fresh Samples, what type of fixation is required for “electron microscopy”?

A

Glutaraldehyde fixation

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13
Q

It is the process of the spooning or scooping the tissue out of the endometrial or cervical canal

A

Curettage

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14
Q

The specimen is washed with __________ to achieve maximum penetration of fixative

A

normal saline

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15
Q

This is the ability to remove the adipose substrate from any specimen and it is important in the study of lymph nodes

A

Fat Clearance

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16
Q

2 groups of Chemical Fixatives, classified according to their mechanism of action

A

Crosslinking Fixative

Precipitating (or denaturing) Fixatives

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17
Q

These are fixatives that are made up of two or more fixatives which been added together to obtain the optimal combined effect

A

Compound Fixative

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17
Q

Formaldehyde/Formalin Derivatives

“Most widely used fixative for routine histology”

Buffer: pH = 7

Best fixative for “Iron pigments and elastic fibers”

A

10% Neutral Buffered Formalin

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18
Q

Examples of Simple Fixatives (AAAAMOP)

A

Aldehydes
Acetic Acid
Acetone
Alcohol
Metallic Fixatives
Osmium
Picric Acid

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19
Q

2 Types of Aldehydes (Cross-Linking Fixative)

A

Formaldehyde
Glutaraldehyde

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19
Q

Most commonly used fixative in histology, which fixes the tissue by forming cross-linkages in the proteins

A

Formaldehyde

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19
Q

Difference between “Formaldehyde” and “Formalin”

A

Formaldehyde: gas produced by oxidation of methyl alcohol

Formalin: made with formaldehyde but the percentage denotes a different formaldehyde concentration

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20
Q

Best fixative for iron pigments and elastic fibers

A

10% Neutral Buffered Formalin

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21
Q

Formaldehyde/Formalin Derivatives

Simple microanatomical fixative made up of “saturated formaldehyde diluted to 10% with sodium chloride”

Fixation Time: 12-24 hours

Preservation of “lipids”, especially phospholipids

Tissues tend to shrink during alcohol dehydration

A

10% Formal-Saline

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22
Q

Identify the Formaldehyde/Formalin Derivative based on their application

  1. Immunohistochemistry and FISH
  2. Fixation of CNS Tissue and post-mortem tissues
  3. Immunohistochemistry only
  4. Routine post-mortem tissues
A
  1. 10% Neutral Buffered Formalin
  2. 10% Formal-Saline
  3. Zinc-Formalin (unbuffered)
  4. Formol-Corrosive (Formol-Sublimate)
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22
Q

Formaldehyde/Formalin derivative best for the preservation of “lipids”, especially phospholipids

A

10% Formal-Saline

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23
Q

Formaldehyde/Formalin Derivatives

Was devised as alternatives to “mercuric chloride” formulations

A

Zinc Formalin (unbuffered)

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23
Q

Identify the Metallic Fixative

Fixation Time: 4-24 hours

“Brown Pigments” produced due to lysis of RBC (if prolonged fixation)

A

Zenker-Formol (Helly’s Solution)

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24
Q

Made up of 2 formaldehyde residues, linked by a three-carbon chain.

It has a larger molecule than formaldehyde, thus the rate of diffusion is “SLOW”

A

Glutaraldehyde

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24
Q

Formaldehyde/Formalin Derivatives

Fixation Time: 3-24 hours

Used in “silver reticulum methods”

Brightens cytoplasmic and metachromatic stains

No need for washing-out

A

Formol-Corrosive (Formol-Sublimate)

24
Q

2 Types of Metallic Fixatives

A

Mercuric Chloride
Chromate Fixatives

25
Q

This fixative leads to formation of “black granular” deposits in the tissues

A

Mercuric Chloride

25
Q

Identify the Metallic Fixative

Fixation Time: 12-24 hours

May act as “mordant”

Stable, but will not be stable when added with acetic acid

A

Zenker’s Solution

25
Q

A metallic fixative used for PTAH Staining, congested specimens, and Trichome Staining

PTAH = Phosphotungstic acid-haematoxylin

A

Zenker’s Solution

25
Q

Most common metallic fixative

Frequently used in saturated aqueous solutions of 5-7%

Penetrates poorly and produces shrinkage of tissues, so it is usually combined with other fixative agents

A

Mercuric Chloride

25
Q

A metallic fixative used for “bone marrow, extramedullary hematopoiesis, and intercalated discs of cardiac muscles”

A

Zenker-Formol (Helly’s Solution)

26
Q

Zenker-Formol is also known as?

A

Helly’s Solution

27
Q

Identify the Metallic Fixative

4% aqueous formaldehyde with 0.22 M mercuric chloride and 0.22 M acetic acid

Enhances nuclear detail

A

Lillie’s B-5 Fixative

28
Q

A metallic fixative used for identifying “normal and abnormal cell types” in bone marrow specimens

A

Lillie’s B-5 Fixative

29
Q

Identify the Metallic Fixative

Fixation Time: 3-12 hours

Excellent cytologic fixative
Penetrates and fixes tissues rapidly and evenly

Tissue should be transferred directly to a high-grade alcohol, to avoid undue swelling of tissues

A

Heidenhains Susa Solution

30
Q

A metallic fixative used for “tumor biopsies” especially of the skin

A

Heidenhain’s Susa Solution

31
Q

A chromate fixative used in 1-2% aqueous solution, usually as a constituent of a compound fixative

precipitates “all proteins” preserves “carbohydrates”

Strong oxidizing agent, hence, a strong reducing agent

A

Chromic Acid

32
Q

A chromate fixative used in a 3% aqueous solution

preserves “lipids” and “mitochondria”

A

Potassium Dichromate

32
Q

Regaud’s Fluid is also known as?

A

Muller’s Fluid

33
Q

Identify the Chromate Fixative

Fixation Time: 12-48 hours

Hardens tissues better and more rapidly than Orth’s Fluid

Demonstration of:
- chromatin
- mitochondria
- mitotic figures
- Golgi bodies
- Rbc
- Colloid-containing tissues

A

Regaud’s Fluid (Muller’s)

34
Q

Identify the Chromate Fixative

Fixation Time: 36-72 hours

Demonstrates “Rickettsiae and other bacteria”

Preserves “myelin” better than buffered formalin

Applied in the study of “early degenerative processes” and “tissue necrosis”

A

Orth’s Fluid

35
Q

An explosive hazard in dry form and normally used in strong saturated aqueous solution

Dyes the tissues “yellow”

A

Picric Acid

36
Q

Penetrates tissue well to react with “histones” and “basic proteins”

Preserves: glycogen
Extracts: lipids

Causes a loss of basophilia unless specimen is thoroughly washed following fixation

A

Picric Acid

37
Q

3 Types of Acetic Acid

A

Glacial Acetic Acid
Lead Fixatives
Trichloroacetic Acid

38
Q

Identify the Picrate

Fixation Time: 4-18 hours

Gives very good results with tissue that is subsequently stained with “trichome”

Preserves: Glycogen
Lyses: RBC
Not used: Frozen sections

A

Bouin’s Solution

39
Q

A picrate applied in fixation of:

Embryos
Pituitary biopsies
GIT biopsies
Animal embryos
Endocrine gland tissues

A

Bouin’s Solution

40
Q

Identify the Picrate

Fixation Time: 4-18 hours

Produces “less lysis” than Bouin’s Solution

Has “decalcifying properties”

For GIT and Endocrine Tissues

A

Hollande’s Solution

41
Q

Identify the Picrate

Fixation Time: 4-18 hours

An alcoholic Bouin’s solution that appears to improve upon aging

Yellow-stain is useful when handling fragmentary biopsies

Not suitable for fixing kidney structures, lipid, and mucus

Produces RBC hemolysis

Preserve: Glycogen and Carbohydrates

A

Gendres Solution

41
Q

Identify the Picrate

Better and less messy than Bouin’s Solution

Fixative for Glycogen

A

Brasil’s Alcoholic Picroformol Fixative

42
Q

Identify the Acetic Acid:

Recommended for “acid mucopolysaccharides”

Fixes: Connective tissue mucin

Used in 4% aqueous solution of basic lead acetate

A

Lead Fixatives

42
Q

Identify the Acetic Acid:

Fixes and precipitates nucleoproteins

Precipitates: Chromosomes and chromatin materials

Causes tissues to swell

Destroys: Mitochondria and Golgi elements

Corrosive to skin

Solidifies at 16C when undiluted

A

Glacial Acetic Acid

43
Q

Identify the Acetic Acid:

Precipitates: proteins and nucleic acids

Marked swelling effect on tissues and used as a weak decalcifying agent

Suitable for small pieces of tissues or bones

A

Trichloroacetic Acid

43
Q

Used to fix specimens at cold temperatures (0-4C = book) (-5 - 4C = ppt)

Not recommended as morphological fixative for tissue blocks

A

Acetone

44
Q

Rapidly denatures and precipitates proteins by destroying hydrogen and other bonds

Uses: 70-100% concentration

A

Alcohol

44
Q

Identify the Alcohol

Fixation Time: 12-18 hours at 3C

Produces better reaction in Feulgen stain than Carnoy’s fluid

Acts both as a “nuclear and histochemical fixative”

Fixes: Mucopolysaccharides and nuclear proteins

A

Newcomer’s Fluid

44
Q

Recommended for the study of “water diffusible enzymes” especially “phosphatases and lipases”

Fixes: brain tissue for diagnosis of rabies

Solvent: For metallic salts to be used in freeze substitution techniques for tissue blocks

Dissolves: Fat
Preserves: Glycogen (poorly)

A

Acetone

45
Q

Identify the Alcohol

Fixes and dehydrates at the same time
Slow penetration

Fixes: Dry and wet smears, blood smears, and BM tissues

A

Methyl Alcohol (100%)

45
Q

Pale-yellow powder which dissolves in water (up to 6% at 20C) to form strong oxidizing solution

Causes “complete denaturation” of protein

1% OsO4 buffered at pH 7.3-7.5 with acetate-veronal buffer is recommended as an appropriate fixative for electron microscopy

A

Osmium

46
Q

Identify the Alcohol

Used for fixing “touch preparations”

Used for special staining procedures such as “Wright-Giemsa”

A

Isopropyl Alcohol (95%)

47
Q

Identify the Alcohol

Fixation Time: 18-24 hours

Preserves but does not fix “glycogen”

Preserves: glycogen, nucleoprotein, nucleic acids

Strong reducing agent

A

Ethyl Alcohol (70-100%)

48
Q

Identify the Alcohol

Fixation Time: 1-3 hours

“Most rapid fixative”

May be used for urgent biopsies

Produces RBC hemolysis, dissolves lipids, and can produce excessive hardening and shrinkage

Fixes: brain tissues for diagnosis of rabies

Used for “curettings”

A

Carnoy’s Fixative

49
Q

Identify the Alcohol

Fixation Time: 12-24 hours

Used during processing to complete fixation

Fixation or post-fixation of “large-fatty” specimens

A

Alcoholic Formalin

49
Q

Identify the Alcohol

Fixation is faster

Used for rapid diagnosis because it fixes and dehydrates at the same time

Fixes: sputum
Frozen section room

A

Gendre’s Fixative

49
Q

Identify the Alcohol

Fixation Time: 3-4 hours

Produces fair results after conventional processing if fixation time is kept very short

Preserves: Nucleic acids
Extracts: Lipids

For frozen sections and smears

A

Clarke’s Solution

49
Q

Identify the Osmium Fixative

“Most common” chrome-osmium acetic acid fixative used

Fixation Time: 24-48 hours

“permanently fixes FAT”

Poor penetrating agent

Excellent fixative for “nuclear structures” (chromosomes)

A

Flemming’s Solution

50
Q

Identify the Alcohol

Fixation Time: 1-6 hours

“Faster acting agent” than alcoholic formalin

Produce “formalin pigment”

A

Formol-Acetic Alcohol

51
Q

Identify the Osmium Fixative

Made up of only “Chromic and Osmic Acid”

Fixation Time: 24-48 hours

“permanently fixes FAT”

Poor penetrating agent

Fixative for “cytoplasmic structures” particularly the “mitochondria”

A

Flemming’s Solution without Acetic Acid

52
Q

Involves thermal coagulation of tissue proteins for rapid diagnosis, usually employed for frozen sections and precipitation of bacteriologic smears.

A

Heat Fixation

53
Q

Type of Fixative that permit the general microscopic study of tissue structures without altering the structural pattern and normal intracellular relationship of the tissues in question

A

Microanatomical Fixatives

54
Q

Microanatomical Fixatives (10-10HFZZBB)

A

● 10% formal saline
● 10% neutral buffered formalin
● Heidenhain ‘s Susa
● Formal sublimate (formal corrosive)
● Zenker ‘s solution
● Zenker-formal (Kelly ‘s solution)
● Bouin’s solution
● Brasil’s solution

55
Q

A type of fixative that preserve specific parts and particular microscopic elements of the cell itself

A

Cytological Fixatives

56
Q

A cytological fixative that preserve the “nuclear structures”

Contain “glacial acetic acid” due to its affinity for nuclear chromatin

pH = 4.6 or less

A

Nuclear Fixative

56
Q

5 Nuclear Fixatives (FCBNH)

A

Flemming’s Fluid
Carnoy’s Fluid
Bouin’s Fluid
Newcomer’s Fluid
Heidenhain’s Susa

57
Q

A cytological fixative that preserve “cytoplasmic structures”

Does NOT contain GAA = destroys the structures

pH = >4.6

A

Cytoplasmic Fixatives

58
Q

5 Cytoplasmic Fixatives (FFORK)

A

Flemming’s without acetic acid
Formalin with “post-chroming”
Orth’s Fluid
Regaud’s Fluid (Muller’s)
Kelly’s Fluid

59
Q

A cytological fixative that preserve the “chemical constituents” of cells and tissues

A

Histochemical Fixatives

60
Q

4 Histochemical Fixatives (FANA)

A

Formal Saline 10%
Absolute Ethyl
Newcomer’s Fluid
Alcohol Acetone

61
Q

To make special staining techniques possible, adding the secondary fixative makes it act as a?

A

Mordant

62
Q

Form of secondary fixation whereby a primarily fixed tissue is placed in aqueous solution of 2.5-3% potassium dichromate for 24 hours to act as a mordant for better staining effects

A

Post- Chromatization

Histopathology Lecture (2 decks)

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