HEMATOXYLIN AND EOSIN (H&E) STAINING

Question 1

REMEMBER!!

H&E is the most widely used histological stain and it is the MOST COMMON STAINING METHOD UTILIZED FOR MICROANATOMICAL STUDIES OF TISSUES using REGRESSIVE type of staining

Question 2

In H&E staining, all fixatives can be used except:

Answer 2

Osmium tetroxide (Osmic acid) because it inhibits hematoxylin

Question 3

What phenomenon indicates that the hematoxylin stain has to be filtered?

Answer 3

A SHEEN of oxidized dye on the surface of hematoxylin solution indicates STAIN HAS TO BE FILTERED

Question 4

Most valuable staining reagent used in histologic work; most widely used powerful NUCLEAR stain; attaches to sulfate groups of DNA stains NUCLEUS

Answer 4

HEMATOXYLIN

Question 5

Most widely used CYTOPLASMIC stain in routine staining sections

Answer 5

EOSIN

Question 6

Three (3) types of EOSIN:

Answer 6

1. EOSIN Y
2. EOSIN S
3. EOSIN B

Question 7

Eosin yellowish, eosin-water-soluble, MOST WIDELY USED type:

Answer 7

Eosin Y

Question 8

Ethyl eosin, eosin alcohol-soluble:

Answer 8

Eosin S

Question 9

Eosin bluish, erythrosin B rarely used

Answer 9

Eosin B

Question 10

Hematoxylin stains the nucleus in color:

Answer 10

Blue to Blue-Black

Question 11

Eosin stains the cytoplasm:

Answer 11

Pink

Question 12

REMEMBER!!

Hematoxylin
- Primary stain
- Basic dye
- Nuclear stain
- STAINS THE NUCLEUS
- STAINS ACIDIC (OR BASOPHILIC) STRUCTURES
- Nucleus: Blue to Blue-Black

Eosin
- Secondary/Counterstain
- Acidic dye
- Cytoplasmic stain
- STAINS THE CYTOPLASM
- STAINS BASIC (OR ACIDOPHILIC) STRUCTURES
- Cytoplasm: Pink

Question 13

COMPOSITION OF H&E STAIN:

Answer 13

1. Fixative
2. Primary stain
3. Differentiator
4. Blueing agent
5. Eosin

Question 14

All fixatives can be used for H&E staining EXCEPT:

Answer 14

FLEMMING’S FLUID because it inhibits hematoxylin

Question 15

Primary stain of H&E stain:

Answer 15

Harris hematoxylin

Question 16

Mordant of Harris hematoxylin:

Answer 16

Potassium alum

Question 17

Differentiator used for H&E stain:

Answer 17

1. Acid alcohol
2. Hydrochloric acid (HCl)

Question 18

MOST IMPORTANT STEP IN REGRESSIVE H&E STAINING

Answer 18

ACID ALCOHOL (DIFFERENTIATOR)

Question 19

Differentiator acid used in combination with alcohol in an acid alcohol solution:

Answer 19

HYDROCHLORIC ACID (HCl)

Question 20

Examples of BLUEING AGENT:

Answer 20

Blueing agents: “SALA”

• Scott’s Tap Water
• Ammonium hydroxide
• Lithium carbonate
• Ammonia water (0.5%)

Question 21

H&E STAIN FORMULA:

DYE:
DYE SOLVENT:
MORDANT:
SOLVENT:
RIPENING AGENT:
ENHANCES NUCLEAR STAINING:

Answer 21

H&E STAIN FORMULA

DYE: Hematoxylin
DYE SOLVENT: Absolute Ethyl Alcohol
MORDANT: Ammonium/Potassium Alum
SOLVENT: Distilled water
RIPENING AGENT: Mercuric oxide
ENHANCES NUCLEAR STAINING: Glacial acetic acid

Question 22

What is the first step in H&E staining that uses xylene?

Answer 22

Deparaffinization

Question 23

In H&E staining, ________ grades of alcohol is used in hydration

Answer 23

DECREASING grades