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HISTOPATH MTAP > HEMATOXYLIN AND EOSIN ROUTINE STAINING > Flashcards

HEMATOXYLIN AND EOSIN ROUTINE STAINING Flashcards

1
Q

● Most widely used histological stain

A

HEMATOXYLIN AND EOSIN ROUTINE STAINING

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2
Q

serves as the basic or primary stain, or nuclear stain (purpose in routine H&E)

A

Hematoxylin

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3
Q

● First person to use paraffin wax for embedding?

A

Butschlii

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3
Q

● Natural dye
○ Derived from extraction from Heartwood of a Mexican tree

A

HEMATOXYLIN

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4
Q

First person to use hematoxylin in histology (1862)

A

Waldeyer

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4
Q

Scientific Name of Mexican tree

A

Hematoxylin campechianum

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5
Q

● May be done by exposing the substance to air and sunlight but the process is SLOW

A

RIPENING/OXIDATION

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6
Q

May be done by adding oxidizing agents such as: (5)

A

○ Hydrogen peroxide
○ Mercuric oxide
○ Potassium permanganate
○ Sodium perborate
○ Sodium iodate

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7
Q

What is the ripening agent of Harry’s hematoxylin?

A

Mercuric Oxide

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8
Q

What is the ripening agent for Erlich’s Hematoxylin?

A

Sodium Iodate

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9
Q

● Used in routine H&E
● Recommended for progressive staining of tissues but it does not mean that it cannot be used for regressive

A

ALUM HEMATOXYLIN

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10
Q

● Mordant: Potash alum
(potassium aluminum sulfate or
simply “alum”)
● Produce good nuclear stai

A

ALUM HEMATOXYLIN

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11
Q

○ Useful general purpose
hematoxylin

A

Harris

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12
Q

Examples of Alum Haematoxylin (7)

A

● Harris
● Ehrlich’s
● Delafield’s
● Gill’s
● Mayer’s
● Cole’s
● Carazzi

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13
Q

Examples of Ripening Agent (6)

A

● Mercuric oxide
● Natural or sodium iodate
● Natural
● Sodium iodate
● Alcoholic iodine
● Potassium iodate

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14
Q

Iron salts are used as oxidizing agents and mordant

A

IRON HEMATOXYLIN

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15
Q

What is the mordant in Harry’s hematoxylin?

a. Mercuric oxide
b. ALUM
c. Both
d. Neither

A

B. Alum

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16
Q

● Mordant is ferric chloride
● In combination with van
Gieson’s stain, can demonstrate
connective tissue elements and
entamoeba histolytica in sections

A

Weigert’s

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17
Q

● Standard iron hematoxylin
● For muscles/connective tissue
fibers

A

Weigert’s

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18
Q

Used for staining elastic fibers

A

Verhoeff/Verhöeff

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18
Q

● Ferric ammonium sulfate
● For mitochondria, muscle
striations, chromatin, and myelin

A

Heidenhain’s Hematoxylin

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19
Q

● Muscle striations
● Mitochondria and myelin

A

Heidenhain’s Hematoxylin

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20
Q

Used for staining myelin

A

Loyez

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21
Q

is an organelle that can be
stained with heidenhain’s hematoxylin but it can also be stained with Janus Green B

A

Mitochondria

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21
Q

is good in demonstrating
collagen

A

Van Gieson’s stain

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22
Q

Chromatin can also be stained with ______________ stain

A

carmine’s

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23
Q

TUNGSTEN HEMATOXYLIN example:

A

Mallory’s PTAH (Phosphotungstic
Acid Hematoxylin)

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24
Q

● To ripen
○ Stand in light for several
weeks or use potassium
of immediate ripening

A

Mallory’s PTAH (Phosphotungstic
Acid Hematoxylin)

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25
Q

● Used for staining
○ Muscle striation
○ Fibrin
○ Glial fiber

A

Mallory’s PTAH (Phosphotungstic
Acid Hematoxylin)

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25
Q

What stain is used to demonstrate muscle Striation?

A

Mallory’s PTAH

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26
Q

Other stains that demonstrate muscle striations?

A

Heidanhain’s Hematoxylin

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27
Q

Used for the study of spermatogenesis

A

COPPER HEMATOXYLIN

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28
Q

MOLYBDENUM HEMATOXYLIN example:

A

Thomas Hematoxylin

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28
Q

What particular part of the male reproductive system does the site of spermatogenesis or the production of sperm cell occur?

A

Seminiferous tubules

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29
Q

Used for staining:
● Collagen
● Endocrine cell granules

A

Thomas Hematoxylin

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30
Q

What other stain presents collagen?

A

Van Gieson Stain

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31
Q

LEAD HEMATOXYLIN example:

A

Solcia Hematoxylin

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32
Q

Used for staining:
● Endocrine cell granules

A

Solcia Hematoxylin

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33
Q

● In the process of H&E staining, the eosin act as a secondary stain, a counterstain, acidic stain, or cytoplasmic stain

A

EOSIN

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34
Q

● Eosin Yellowish
● Most commonly used

A

Eosin Y

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35
Q

● Eosin Bluish
● Produces a deeper red color

A

Eosin B

36
Q

Ethyl Eosin

A

Eosin S

37
Q

■ Temperature to fix surgical specimens (room temp)
■ Formaldehyde for 24hrs

A

Fixation

38
Q

■ Most common dehydrating agent is alcohols
■ Should be used in ascending rates
■ We intend to infiltrate the paraffin wax unto the tissue but it is not soluble to water nor to alcohol, this we need to remove alcohol

A

Dehydration

39
Q

■ Otherwise known as dealcoholization.
■ Makes the tissue transparent and clears the alcohol
■ Usually used is Xylene
● Which is usually miscible to
paraffin wax

A

Clearing

40
Q

■ Temperature of paraffin oven is 55 - 60 °C or 5°C above the melting point of the wax.

A

Impregnation

41
Q

■ We get the tissue from the melted paraffin wax, open the tissue cassette and put it in a mold to perform embedding
■ Also known as casting
■ Temp of paraffin wax is 5°C - 10°C above the melting point of the paraffin wax
■ Let the paraffin wax harden

A

Embedding

42
Q

■ Also called as cutting or microtomy
■ Most commonly used is the rotary microtome
● Old sources: 4-6 micron of
tissue
● New sources: 3-5 micron of
tissue
○ Remember both old and
new references, choose
the closest. Depends on
what book the board
examiner uses

A

Sectioning

42
Q

■ Remove the excess paraffin wax
■ Ideal shape is truncated pyramid or 4-sided prism

A

Trimming

43
Q

■ Flotation water bath
● Optimal temperature is 6°C -
10°C below the melting point of
paraffin wax
■ Get a slide and apply Meyer’s Egg Albumin which is composed of;
● Egg white
● Glycerol
● Crystal sulfide (prevent growth
of mold)
■ Fish out, and place it in the paraffin oven to remove the excess paraffin wax
(deparaffinization)
■ Put it in xylene or xylol

A

Sectioning

44
Q

CHANGES OF XYLENE OR XYLOL

○ We need to add xylene to remove the paraffin in the tissue, heat alone could not remove the
paraffin wax

A

Process of deparaffinization

45
Q

The tissue needed water for the stain to adhere

A

Rehydration

46
Q

DESCENDING GRADES OF ALCOHOL

Slowly adding water to the tissue

A

● Rehydration

47
Q

When do we remove fixative artifacts and fixative treatments?

A

It is removed after rehydration and right before primary staining

**this artifacts can obscure the the tissue

48
Q

STAIN WITH HARRIS/ EHRLICH’S/DELAFIELD’S
HEMATOXYLIN

● Primary stains
○ Alum hematoxylin, serve as the primary, basic and nuclear stain of the tissue.
○ The color of the nucleus is already ___________________
○ While the cytoplasm is still __________

A

● light transparent blue

● colored

49
Q

ACID ALCOHOL (DIFFERENTIATOR)
● Decolorizer
○ Differentiation phase
○ The nucleus will turn _____
○ THE cytoplasm would be ______________

A

● RED
● COLORLESS

50
Q

Blueing agents (3)

A

○ Ammonium hydroxide
○ Lithium carbonate
○ Scott’s tap water

51
Q

AMMONIA WATER (AMMONIUM HYDROXIDE, LITHIUM CARBONATE, SCOTT’S TAP WATER)

○ Transform the color of the nucleus back to _____, Due to the secondary stain being being eosin Y, which has a pink color, and this, for differentiation, different colored stains are used in the cell (contrast)

● Cytoplasm: _______________

A

● BLUE
● COLORLESS

52
Q

What is the expected color of the nucleus of the cell if the medtech forgot to use AMMONIA water?

A

RED

53
Q

Eosin Y is called as (4)

A

○ Secondary stain
○ Acidic stain
○ Cytoplasmic stain
○ Counter stain

54
Q

Stain with Eosin Y
● Color of Nucleus: _________
● Color of Cytoplasm: _______

A

● Color of Nucleus: still color BLUE
● Color of Cytoplasm: PALE PINK

55
Q

● Ascending Grades of Alcohol: _____________
● Descending Grades of Alcohol: _____________

A

● Ascending Grades of Alcohol: DEHYDRATION
● Descending Grades of Alcohol: REHYDRATION

55
Q

Known as CLEARING (Dealcoholization)

A

Xylol/ Xylene

56
Q

Purpose of Xylol in the FIRST STEP of routine H&E:

A

DEPARAFFINIZATION

57
Q

Purpose of Xylol in the LAST STEP of routine H&E:

A

CLEARING AGENT

58
Q

Nuclei

A

Blue to Blue Black

58
Q

Karyosome

A

Dark Blue

59
Q

Cytoplasm, proteins in edema fluid

A

Pale Pink

60
Q

Calcium and calcified bone

A

Purplish Blue

61
Q

Muscle fibers

A

Deep Pink

62
Q

● Considered as the staining method of choice for Exfoliative Cytology

A

PAPANICOLAOU STAINING (PAPS STAINING)

63
Q

● aka Harris Hematoxylin
● Serve as nuclear stain
● Gives color to the nucleus
whatever kind of cell it is.
● stain for the nucleus of mature
superficial cells

A

Hematoxylin

63
Q

PAPANICOLAOU STAINING (PAPS STAINING)

Fixative used:

A

95% Ethanol

64
Q

PAPANICOLAOU STAINING (PAPS STAINING)

Three (3) Stains applied:

A

○ Hematoxylin
○ OG6
○ EA

65
Q

● stain for the cytoplasm of the
MATURE superficial cells.

A

OG6 (Orange Green)

65
Q

● stain for the cytoplasm of
IMMATURE vaginal cells.
○ Such as parabasal and
intermediate cells

A

EA 36 or 50 (Eosin Azure)

66
Q

used for staining hemoglobin

A

Benzidine

66
Q

● DNA (green fluorescence)
● RNA (red fluorescence)

A

Acridine Orange

66
Q

Components of EA 36 or 50: (3)

A

○ Eosin Y
○ Light Green SF
○ Brismark Brown

67
Q

● for staining amyloid in frozen
sections and platelets in blood

A

Crystal violet

68
Q

● formed by the mixture of crystal violet, methyl violet, and dexterin

A

Gentian violet

69
Q

● Gold standard for amyloid
demonstration
● stain for axis cylinders in
embryos

A

Congo Red

70
Q

● used as a 4% aqueous solution
in Krajian’s method of staining
elastic tissues, amyloid, and
myelin

A

Congo Red

71
Q

● Oldest of all stains
● stains amyloid, cellulose, starch,
carotenes, and glycogen
● widely used for removal of
mercuric fixative pigments

A

Iodine

71
Q

● contrast stain for staining Ascaris eggs and erythrocytes
● used also as a bacterial spore
stain

A

Malachite Green

72
Q

● used for demonstrating
mitochondria during intravital
staining (or supravital?)

A

Janus Green B

73
Q

● used for demonstration of
neuroglia in frozen sections

A

Victoria Blue

74
Q

● used as a substitute for carbol
fuchsin in acid-fast staining

A

Night Blue

75
Q

● NOT real dyes
● They do NOT have auxochrome
groups.
● They give color to lipids simply
because they are more soluble in
lipid medium of the tissues than
in their medium of 70% alcohol.

A

Lysochromes (Oil Soluble
Dyes)

76
Q

● Examples of oil soluble dyes
used for demonstration of
intracellular fats:
○ Sudan Black B
○ Sudan III
○ Sudan IV (Scharlach R)

A

Lysochromes (Oil Soluble
Dyes)

77
Q

● PAS-positive substances =
red/magenta red
● Mucoproteins are the most
common PAS-positive
substances

A

Periodic Acid-Schiff Reaction

78
Q

● Considered as the most reliable
and specific histochemical stain
for DNA

A

Fill Gel Technique

79
Q

● Can observe Howell-Jolly Bodies
● Howell-Jolly Bodies: Fill Gel
Reaction (+)

A

Fill Gel Technique

80
Q

● Best’s Carmine
● PAS (periodic acid-Schiff)
● Langhan’s Iodine Stain

A

Stains For Glycogen

80
Q

● Used for the demonstration of
spirochetes.

A

Warthin–Starry Stain

81
Q

● Applicable not only for muscle
striations but also for Fibrin
Phosphatase.

A

Mallory’s PTAH (Phosphotungstic
Acid-Hematoxylin)

82
Q

Expected color:
○ Early fibrin: YELLOW
○ Old fibrin: BLUE

A

Lendrum’s MSB (Martius, Scarlet,
Blue)

83
Q

Expected color:
○ Early fibrin:

A

YELLOW

84
Q

Expected color:
○ Old fibrin:

A

Blue

85
Q

PAS-positive substances =

A

Red/Magenta Red

HISTOPATH MTAP (5 decks)

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