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Cell Pathology > L.6 Histochemistry > Flashcards

L.6 Histochemistry Flashcards

1
Q

What is histochemistry?

A

The study of tissue-chemical interactions involved in the visualization of cell/tissue components

Histochemistry aims to create stable, coloured compounds at the site of components to be detected.

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2
Q

What is the aim of histochemical methods?

A

Formation of stable, coloured compounds at the site of the component to be detected with clear contrast to other elements present in tissue

This is essential for observing cellular details.

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3
Q

Why do we need colour to create contrast in histochemistry?

A

Processed tissue and tissue sections are generally colourless and need to be stained to observe cell and tissue detail, components and structure

Various coloured dyes and stains can be used to identify different components.

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4
Q

What are the key staining methods in histochemistry?

A
  1. Staining by direct binding
  2. Metal impregnation
  3. Lysochrome staining
  4. Histochemical
  5. Indirect Staining methods
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5
Q

What is staining by direct binding?

A

Bonding, Affinity for chromogen (direct and indirect) and dye-staining

This method involves directly binding dyes to the tissue components.

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6
Q

What does metal impregnation involve?

A

Metallic Ion staining (silver and gold) and staining by adsorption

This method uses metals to visualize cell structures.

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7
Q

What is lysochrome staining?

A

Staining by solubility and lipid staining (with dyes)

It targets lipids in tissues.

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8
Q

What is histochemical staining?

A

Staining by chemical reaction that forms an insoluble precipitate due to substrate creating a chromogen/coloured compound at the site of reactivity

This reaction is similar to those seen in a test tube.

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9
Q

What are indirect staining methods?

A

Through antibody or oligonucleotide probe binding to substrate and linking to a chromogen (label)

Immunohistochemistry is a common example.

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10
Q

What does acidophilic refer to in histochemistry?

A

Binds to acidic groups/dyes

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11
Q

What does basophilic refer to in histochemistry?

A

Binds to basic groups/dyes

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12
Q

What are the majority of dyes made from today?

A

Man-made sources

Originally, only natural sources were available for dyes.

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13
Q

From which industry are synthetic dyes primarily derived?

A

Petrochemical industry

Specifically, derivatives of benzene.

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14
Q

What are the three possible charges of dyes in an aqueous solution?

A
  • Acidic
  • Basic
  • Neutral
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15
Q

What are the two modes of staining mentioned?

A
  • Progressive staining
  • Regressive staining
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16
Q

What is a characteristic of fluorescent dyes?

A

Emit light under UV

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17
Q

What does metachromatic mean in the context of dyes?

A

Two colours – monomeric and polymeric

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18
Q

What type of dyes are lysochrome?

A

Hydrophobic dyes

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19
Q

What do lysochrome dyes specifically stain?

A

Lipids

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20
Q

What is a common structural feature of all dyes?

A

Aromatic structure

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21
Q

In which range do all dyes absorb light?

A

Visible range of the Electromagnetic spectrum

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22
Q

What is the purpose of the chromophoric structure in dyes?

A

To enable visibility

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23
Q

What is an auxochrome?

A

An added reactive group that aids ionization in water

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24
Q

What does the Colour Index Number identify?

A

Chemical structure, reactivity, and common names of dye

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25
Is the Colour Index Number an internationally accepted system?
True
26
Provide an example of a dye and its Colour Index Number.
Acid Fuchsin CI42685 aka acid violet 19, acid magenta
27
What are mordants in the context of dyeing?
Metal ions that complex with dye to enhance reactivity
28
Name one dye that requires a mordant.
* Haematoxylin * Alcian blue * Celestine blue
29
What is affinity-based staining?
Affinity-based staining relies on specific chemical interactions between reactive groups present in tissue components and those in dyes, metals, or other reagents
30
What is the mechanism of affinity-based staining?
Dyes possess functional groups (e.g. acidic or basic) that bind selectively to opposite-charged tissue molecules
31
Give an example of affinity-based staining.
Masson’s Trichrome Stain
32
What does the light green dye in Masson’s Trichrome Stain specifically bind to?
Basic proteins in collagen
33
What colors are used to differentiate various tissue components in Masson’s Trichrome Stain?
* Collagen: green or blue * Cytoplasm: red * Nuclei: black or dark blue
34
What are metallic impregnation methods used for?
Visualizing delicate structures like nerve fibers, basement membranes, and reticulin
35
What is the first key step in metallic impregnation?
Sensitization
36
What occurs during the impregnation step of metallic impregnation?
A metal salt solution is applied, and tissue structures adsorb the metal ions
37
What is the purpose of the reduction step in metallic impregnation?
To chemically reduce metal ions to their elemental (metallic) state, forming dark, insoluble deposits
38
What does the stabilization (toning) step do in metallic impregnation?
Enhances contrast and stabilizes the metal deposit (e.g., with gold chloride)
39
What is the argentaffin reaction?
Tissue components both bind and reduce silver ions without an external reducer
40
Give examples of tissue components that exhibit the argentaffin reaction.
* Melanin * Lipofuscin * Certain endocrine cells
41
What is the argyrophilic reaction?
Tissue binds silver ions but requires an external reducing agent to complete the reaction
42
Give examples of tissue components that exhibit the argyrophilic reaction.
* Reticulin fibers * Basement membranes * Glycogen * Nerve fibers
43
What mnemonic can help remember the argyrophilic reaction?
aRgyRophillic = Requires Reducer
44
What are lysochrome stains used for?
Demonstrating lipids in tissue sections, especially neutral fats and phospholipids. ## Footnote Lysochrome stains are fat-soluble dyes that highlight specific lipid components in biological samples.
45
How do lysochrome stains work?
They work via hydrophobic interactions and solubility differences rather than chemical bonds. ## Footnote This mechanism allows the dye to selectively color lipid droplets in tissues.
46
What is the mechanism of lysochrome staining?
The dye is more soluble in lipids than in the aqueous or alcoholic solvent, dissolving into lipid droplets. ## Footnote This selective solubility is key to the staining process.
47
Name a common dye that stains neutral fats red.
Oil Red O ## Footnote Oil Red O is widely used in histology for visualizing neutral fats.
48
What color does Sudan Black B stain lipids?
Blue-black ## Footnote Sudan Black B is another important dye used in lipid staining.
49
What does Nile Blue Sulphate differentiate?
Neutral lipids (pink/red) from phospholipids (blue). ## Footnote This differentiation is useful for understanding lipid composition.
50
When are lysochrome stains typically used?
On frozen sections, as lipids are dissolved during standard paraffin processing. ## Footnote Frozen sections preserve lipids, making them suitable for lysochrome staining.
51
What does histochemistry involve?
The use of chemical reactions directly on tissue sections to detect specific biochemical substances. ## Footnote Histochemistry is essential for visualizing various biomolecules in tissues.
52
What are the key criteria for a reliable histochemical technique?
* Sensitivity and Specificity * Compatibility with Tissue Sections * Preservation of Tissue Integrity * Stable and Visible End Product * Site-Specific Reaction * Rapid Reaction Time ## Footnote These criteria ensure that histochemical methods provide accurate and useful results.
53
What is meant by sensitivity and specificity in histochemical techniques?
The method must detect small amounts of the target substance without cross-reacting with unrelated compounds. ## Footnote Sensitivity and specificity are crucial for accurate diagnosis.
54
What must occur for histochemical reactions to be compatible with tissue sections?
The reaction must occur within thin sections of fixed tissue mounted on slides. ## Footnote This compatibility is essential for effective visualization.
55
What is required for the preservation of tissue integrity in histochemistry?
The procedure should not damage or alter the overall tissue morphology. ## Footnote Maintaining tissue integrity is vital for accurate analysis.
56
What characteristics should the final reaction product of a histochemical technique have?
* Insoluble in solvents used during processing * Clearly visible under light microscopy * Durable for long-term storage ## Footnote These characteristics ensure that the results are reliable and maintainable.
57
What does site-specific reaction mean in histochemistry?
The product must precisely localize to the target substance, avoiding background staining. ## Footnote Site specificity enhances the clarity of results.
58
Why is rapid reaction time important in histochemistry?
Reactions should be fast enough to prevent diffusion of the substance from its original location in the tissue. ## Footnote Quick reactions help maintain the spatial accuracy of the staining.
59
What is the purpose of the Perls Prussian Blue Stain?
Detection of ferric (Fe³⁺) iron, typically stored as hemosiderin in tissues ## Footnote Common tissues include liver, spleen, and bone marrow.
60
What is the first step in the Perls Prussian Blue Stain procedure?
Treat with 2% Hydrochloric acid ## Footnote This step liberates ferric iron from protein complexes.
61
What occurs during the second step of the Perls Prussian Blue Stain?
Treat with Potassium Ferrocyanide ## Footnote Ferrocyanide ions complex with ferric iron to form a blue compound (ferriferrocyanide).
62
What does blue staining in the Perls Prussian Blue Stain indicate?
Iron accumulation ## Footnote Useful in diagnosing hemochromatosis, siderosis, or prior hemorrhage.
63
What is the purpose of the Periodic Acid Schiff reaction?
Highlights polysaccharides, mucosubstances, basement membranes, glycogen, and fungal walls.
64
What is the first step in the Periodic Acid Schiff reaction?
Treat with 1% Periodic acid ## Footnote This step oxidizes diglycol groups in hexose molecules to dialdehydes.
65
What happens during the second step of the Periodic Acid Schiff reaction?
Treat with Schiff reagent ## Footnote Dialdehydes bind and recolour basic fuchsin (magenta).
66
What does magenta staining in the Periodic Acid Schiff reaction indicate?
Presence of glycogen, mucin, basement membranes, or fungal elements.

Cell Pathology (28 decks)

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