Part 1 Flashcards by Janice Mariel Bassig

Enumerate the tissue processing

Fixation
Dehydration
Clearing
Impregnation
Embedding
Trimming
Section cutting
Staining
Mounting
Labeling

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First and most critical step in tissue processing

Fixation

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1’ aim of fixation

Preserve cell

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2’ aim of fixation

Harden and protect tissues

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Different fixatives

Microanatomical fixatives
Cytological
Nuclear
Cytoplasmic
Histochemical

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Formaldehyde stock ang working

Stock: 37-40%
Working: 10% (unstable)

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Best general tissue fixative

10% Neutral Buffered Formalin

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Best fixative for tissue

10% Neutral Buffered Formalin

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Rate of tissue penetration

1mm/hr

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Ratio of FIXATIVE TO TISSUE

20:1

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Ratio of FIXATIVE TO TISSUE (PROLONGED)

50-100:1

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Ratio of DECALCIFYING AGENT TO TISSUE

20:1

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Decalcifying agents

Acids
Chelating agents
Ion exchange resins
Electrophoresis

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Most common decalcifying agent

HNO3 (NITRIC ACID)

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Both FIXATIVE and DECALCIFYING AGENT

5% Formic Acid

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Best general decalcifying agent

5% Formic Acid

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Decalcifying agent for small bones and teeth

5% Formic Acid

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Tissue Softeners

4% phenol
Molliflex (soapy and swollen tissue)
2% Hcl
1% Hcl with 70% alcohol

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Aim of DEHYDRATION

Remove fixatives and water

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Dehydration:
A. Ascending grades of alcohol
B. Descending grades of alcohol

A. Ascending grades of alcohol

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Ratio of DEHYDRATING AGENT to TISSUE

10:1

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Dest dehydrating agent

Ethanol

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Dehydrating agent for BLOOD and TISSUE FILMS

Methanol

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Dehydrating agent for PLANTS and ANIMALS

Butanol

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Denatured Alcohol

ETHANOL and METHANOL

Both FIXATIVE and DEHYDRATING AGENT

Acetone

Both DEHYDRATING AGENT and CLEARING AGENT

Dioxane (diethylene dioxide)

Most commonly used clearing agent

Xylene

Time (CLEARING)

1/2 to 1 hour

Substitute for Xylene/benzene

Toluene

Ratio of INFILTRATING MEDIUM to TISSUE

25:1

Different types of medium

Paraffin wax Celloidin Gelatin Plastic

Paraffin is Introduced by

Butschlii

This medium is not recommended for fatty tissues

Paraffin

Temp (EMBEDDING)

2-5’C above

Double embedding is for?

Brain

Double Embedding 1st: 2nd:

1st: celloidin 2nd: paraffin

Cold knife procedure Knife: Tissue: Environment:

Knife: -40 to -60 ‘C Tissue: 5 to -10’C Environment: 0 to -10’C

Cryostat Temperature

-18 to -20 ‘C

Freeze drying Quenching: Sublimation:

Quenching: -160’C Sublimation: -40’C

Algor mortis (Temp)

7’F/hr 3.89’C/hr

Removing excess WX after embedding

Trimming

Ideal shape for trimming

Four-sided prism

Microtomy ROUTINE HISTOPATH (rotary)

4-6um

Microtomy Freezing

10-15 um

Microtomy Ultrathin

0.5 um

Trefall/Simplest Microtome

Rocking/Cambridge Microtome

Minot MOST COMMONLY USED for Paraffin Embedded Tissues

Rotary/Minot Microtome

Most dangerous microtome

Sliding microtome

Movable knife microtome

Sliding microtome

2 types of Sliding Microtome

Base sledge Standard sliding

Knife to tissue block angle

0-15’ angle

Bevel angle

27-32

Removal of gross nicks

Honing

Heal to toe

Honing

Types of honing

Belgium yellow Arkansas Fine carborundum

Removal of gross burrs

Stropping

Toe to heel

Stropping

Natural Dyes are derived from

Plants and animals

Natural Dyes

Hematoxylin Cochineal dyes: female Coccus cacti Orcein: from lochens Saffron

Aka coal tar dyes

Synthetic dye

Synthetic dye is derived from

Benzene

Color bearer

Chromophores

Dyeing property

Auxochrome

Benze + Chromophore

Chromogen

Chromogen + auxochrome

Dye

Imparts color temporarily

Chromogen

Imparts color permanently

Dye

Hematoxylin other name

Hematoxylin capechianum

Nucler/basic/primary stain

Hemtoxylin

1st to use hematoxylin

Waldeyer

Active coloring substance

Hematein

H & E Staining Steps

1. Xylol 2. Descending grades of alcohol 3. H2O 4. Remove fixative artifact pigments after rehydration and before staining 5. Stain (nucleus: light blue) 6. H2O 7. Acid Alcohol 8. Ammonia water 9. Wash 10. Stain: eosin 11. Ascending grades of alcohol 12. Xylene 13. Mount and label

Oldest stain

Iodine

Stains Hgb

Benzidine

Stains RNA and DNA

Acridine orange

Stains tissue, amyloid, myelin

Congo red

Stains ascaris egg

Malachite green

Stains mitochondria

Janus green

Different types of adhesives

Mayer’s egg albumin Dried albumin Gelatin Gelatin - formaldehyde Starch paste Plasma Poly - L - Lysine

Refractive index of glass

1.518

Contains xylene

Resinous media

For lipids (no xylene)

Aqueous media

Standard mounting medium For fat stains

Glycerin jelly

Canada balsam aka

Abus balsamea

1st to describe the 4 signs of inflammation

Cornelius Celsus

Methods of staining

Direct staining (w/o mordant) Indirect Staining (w/ mordant)

Part 1 Flashcards

(87 cards)