Staining Nuclear, Intro to Surg Path

Staining provides what with light microscopy?

contrast

Disruption of normal cellular patterns are caused by:

Infection

Tumor

What part of the nucleus the stainable component?

Heterochromatin

Heterochromatin is what kind of protein?

tightly wound

What portion of the nucleus is non-stainable?

Euchromatin

Euchromatin is what kind of protein?

loosely bound

The nucleolus can be stained by what stain due to the dense protein content?

Eosin

Hemotoxylin stains acidic structures:

blue

Eostin stains basic structures:

pink

The eosin will stain what portion of the cell?

cytoplasm

the hemotoxylin will stain what portion of the cell?

nucleus

Covalent bonds share electrons and are:

strong

Ionic bonds are positive attracted to negative and are:

strong

Hydrogen bonds are:

covalently H+ attracted to strong electro-negative charge, weak

Van der Waal bonds are:

electrostatic charge and are very weak

Physical staining is purely by:

ABsorption

What occurs in physical staining?

Passing the dye molecule from the solution to the tissue component to be demonstrated. the component stained becomes the same color as the dye solution

Chemical staining occurs by:

ADsorption

H and E staining uses what kind of staining?

Adsorption

What occurs in an H and E stain?

A chemical union or reaction between a dye and stained substances. it occurs by ions within stain solutions being attracted to and influenced by ions in tissues. Dye molecules precipitate on certain cellular components

What is metachromatic?

A change in color

A single dye reacts with tissue cells and stains them different colors

What is polychromatic?

Contains or more dyes

dye solutions stain tissue components different colors by a phenomenon other than metachromasia

In absorption, molecules are:

draw INTO the material

In adsorption, molecules:

adhere to surface

What are some examples of stains that use adsorption?

IHC

Diff Quik

Eosin is acidic and has a negative charge (anionic dye), so it is attracted to:

basic (+) cytoplasmic proteins

Cytoplasmic granules

connective tissue

red blood cells

Nucleolus

Hemotoxylin is basic and has a positive charge (cationic dye), so it is attracted to:

acidic (-) nucleus

heterochromatin

ER

Acidophilic means:

tissues attracting acid dyes (anionic)

Basophilic means:

Tissues attracting basic dyes (cationic)

Acid and Basic dyes have little to do with pH but rather:

the charge

The nucleus is negatively charged, so it attracts:

basic dyes

The cytoplasm is positively charged, so it attracts:

acidic dyes

What is IEP (isoelectric point)?

Point where positive and negative charges are equal

No migration between charged proteins at what pH?

6

What is ideal cytoplasmic staining?

pH must be between pH 4-5

If the pH is above 6, what occurs?

Proteins have net - charge, won’t attract anionic (-) dyes like eosin

Eosin will not attach to cytoplasmic protein

Pale to no staining due to net negative charge

If the pH is below 6, what occurs?

Proteins have a net (+) charge, attract anionic (-) dyes like eosin

the darkest portion of the cytoplasm that is dyed by eosin should be:

Erythrocytes

The intermediate portion of the cytoplasm that is dyed by eosin should be:

Collagen, muscle

the lightest portion of the cytoplasm that is dyed by eosin should be:

cytoplasm

What does pholxine do?

Added to give more vibrant colors

Hematoxylin comes from what?

Nuclear dye from the logwood tree (Haematoxylon campechianum)

Can hematoxilyn be used alone as a dye?

No, it must be oxidized

What are some examples of a metal mordant?

Aluminum

Iron

Chromium

In order to use hematoxylin, what must occur?

Oxidize

use metal mordant as a link

Create a dye lake

What part of the cell is basophilic?

Nucleus

What part of the cell is acidophilic at the proper pH?

Cytoplasm, collagen

What is the proper pH for acid stains?

4-5 pH

Cytoplasm attracts anionic or cationic proper pH?

Cationic

What is the typical thickness of a glass slide?

1.0-1.5 mm

What is the function of coverglass?

Used for preservation of stained tissue sections

Provide better microscopic magnification and examination

Improved refractive index

How long should slides be allowed to dry before filing?

24 hours

Staining issues could be caused from:

Fixation or processing

Water that contains chlorine, sulfur, or iron, which changes the pH (USE DISTILLED WATER)

What could occur if slides dry out during staining before coverslipping?

Cornflaking

Why should solutions be covered when not in use?

Prevents atmospheric moisture from getting into alcohol

Prevents evaporation

Routine H and E staining:

Xylene (remove paraffin from slide)

Graded alcohols (hydrate)

Water (rinse)

Hematoxylin (nuclear stain, now purple color)
Water (remove excess hematoxylin)

Clarifier (acid alcohol, clear hematoxylin from cytoplasm

Water (rinse clarifier from slide)

Bluing (blue nuclei)

Water (remove excess bluing from slide)

Alcohol (prevents eosin from getting diluted with water)

Eosin (cytoplasmic stain)

Xylene (clear)

Mounting media

What do charged microscope slides do?

To anchor tissue to the slide electrostatic attraction

What is a dye lake?

Stable coordination complex formed by combining a mordant with a dye molecule (like oxidized hematoxylin) which then binds tissue components