FIXATION

Question 1

Defined as the killing, penetration, and hardening of tissues

First and most critical step in tissue processing

Answer 1

Fixation

Question 2

Fixation

Primary purpose:

Answer 2

Preserve the morphologic and chemical integrity of the cell in a life-like manner as possible

Question 3

EFFECTS OF FIXATIVES

•______ soft tissues in preparation for further tissue processing

• Render cells resistant to____ caused by chemicals used in further processing

• Inhibit_____ caused by bacteria and fungi

• Minimize the risk of_____

• Act as_____ for certain stains, thus promoting or hastening staining, or inhibit certain dyes

• Reduce the risk of infections during handling and actual processing of tissues

Answer 3

Hardens

damage

decomposition

occupational infection

mordant

Question 4

CHARACTERISTICS OF A GOOD FIXATIVE

Answer 4

• Cheap
• Stable
• Safe
• Quick
• Inhibits bacterial decomposition
• Produce minimum shrinkage
• Rapid and even penetration
• Hardens the tissue
• Makes cellular contents resistant to further processing
• Permit staining

Question 5

FACTORS AFFECTING FIXATION

Answer 5

• Fixative of Choice
• Time
• Tissue to fixative ratio
• Penetration rate
• Thickness of specimen
• Tissue components
• pH
• Temperature
• Osmolality
• Agitation, vacuum

Question 6

Fixative of Choice

Morphologic criteria for dx have been established based on
Formalin-Fixed Paraffin
Embedded Specimen (FFPES)

Answer 6

10% Neutral Buffered Formalin

Question 7

Time
Must be performed as soon as possible;
_________ after blood supply is cut off

Answer 7

20-30 mins

Question 8

Tissue-to-Fixative Ratio
•_______(tissue to fixative ratio)

Answer 8

1:10 or 1:20

Question 9

Penetration Rate
Formalin:_____(but slows down as it goes deeper into the tissue)

Answer 9

1mm/hr

Question 10

Thickness of Specimen
• Larger -› Longer fixation time, more fixative

• Light Microscopy:
• Electron Microscopy:

Answer 10

2cm2 x 0.4cm

1-2mm2

Question 11

Tissue Components
• Longer Fixation time:

Answer 11

• Fibrous Tissue
• Mucus
• Fat
• Blood

Question 12

Tissue Components

Shorter Fixation time:

Answer 12

Small of loosely textured tissues

Question 13

рН

Optimal pH:
Use buffers

Answer 13

6 to 8

Question 14

pH

For Electron Microscopy:

Answer 14

pH should match physiologic pH

Question 15

• Temperature
Higher temp ->

Answer 15

faster fixation rate and autolysis

Question 16

Optimal temp

Answer 16

Room temp to 45C

Question 17

Tissue processor temp

Answer 17

40C

Question 18

Microwave processing temp

Answer 18

Up to 65C

Question 19

Electron microscopy temp

Answer 19

0-4C

Question 20

Tuberculosis temp

Answer 20

100C

Question 21

Rapid biopsy temp

Answer 21

60C

Question 22

Osmolality

Hypertonicity ->

Isotonicity and Hypotonicity -›

Answer 22

cell shrinkage

cell swelling

Question 23

Osmolality

maintain tissues at

Answer 23

slightly hypertonic solution (400-450 mOsm)

Question 24

TYPES OF FIXATIVE
• Based on Composition

Answer 24

Simple
Compound

Question 25

______ Made of only one component ______ Consists of two or more components of fixatives

Answer 25

Simple Compound

Question 26

TYPES OF FIXATIVE BASED ON ACTION

Answer 26

Microanatomical Cytological Histochemical

Question 27

General study of tissues w/o structure alteration

Answer 27

Microanatomical

Question 28

• Cytological ________ _______

Answer 28

• Nuclear • Cytoplasmic

Question 29

Nuclear •_____ Glacial acetic acid has affinity to nuclear chromatin Cytoplasmic •_____ • HAc destroys mitochondria and Golgi bodies

Answer 29

pH ≤ 4-6 pH > 4-6

Question 30

• Preserves chemical constituents if cells and tissues

Answer 30

Histochemical

Question 31

Microanatomical

Answer 31

-10% NBF - 10% Formol-Saline - Heidenhain's Susa - Zenker's - Zenker-formol (Helly's) - Bouin's - Brail's

Question 32

Nuclear

Answer 32

-Flemming's with glacial acetic acid - Carnov's - Bouin's - Newcomer's - Heidenhain's

Question 33

Cytoplasmic

Answer 33

- Helly's - Orith's - Regaud's / Molter's - Formalin with Post-charming -Fleming's w/o glacial acetic acid

Question 34

Histochemical

Answer 34

- 10% Formol Saline - Absolute ethanol - Newcomer's - Acetone

Question 35

ALDEHYDE FIXATIVES

Answer 35

1. Formaldehyde 2. 10% formol saline 3. 10% Neutral Buffered Formalin (NBF) 4. Formol-Corrosive (Formol Mercuric Chloride) 5. Gendre's (Alcoholic Formalin) 6. Hollande's 7. Glutaraldehyde 8. Paraformaldehyde 9. Karnovsky's Paraformaldehyde / Glutaraldehyde 10. 40% Aqueous Glyoxal

Question 36

3 types of metallic fixatives

Answer 36

Mercuric chloride Chromates Lead

Question 37

Mercuric Chloride

Answer 37

Zenker Zenker-Formol (Hellv's) Carnoy-Lebron [Heidenhain's Susa B5 Schaudinn's

Question 38

Chromates

Answer 38

Chromic acid [Regaurd's/Muller's 'Orth's Potassium dichromate

Question 39

PICRIC ACID FIXATIVES

Answer 39

1. Bouin’s 2. Brasil's Alcoholic Picroformol 3. Glacial Acetic Acid

Question 40

ALCOHOL FIXATIVES

Answer 40

1. Carnoy's Fluid 2. Ethanol (70 - 100%) 3. Methanol / Wood Alcohol (100%) 4. Isopropanol 5. Newcomer's Fluid 6. Gendre's (Alcoholic Formalin)

Question 41

OSMIC ACID FIXATIVES

Answer 41

1. Flemming's Solution w/ GAC 2. Flemming's Solution w/o GAC 3. Trichloroacetic Acid 4. Acetone