FINALS - 1 - Rapid Tissue Processing

Question 1

Advantage of rapid tissue processing

Answer 1

Faster and safer (no formalin or xylene)

Question 2

Conventional vs Rapid tissue processing

Answer 2

Conventional = longer process; Rapid = shorter (4 steps)

Question 3

Microwave role in tissue processing

Answer 3

Speeds up processing by heat penetration

Question 4

Why microwave processing is fast

Answer 4

Uses heated alcohols; increased molecular movement

Question 5

Microwave processing time

Answer 5

2.5 hours (60–80% faster)

Question 6

Steps in microwave processing

Answer 6

1 ethanol, 1 isopropanol, 2 paraffin wax (4 total)

Question 7

Can you use a household microwave?

Answer 7

No – lacks temp control, stirrer, probe

Question 8

Magnetic stirrer function

Answer 8

Makes irradiation even

Question 9

Temperature probe use

Answer 9

Maintains precise temperature

Question 10

Microwave staining advantages

Answer 10

Shorter time, better contrast, less non-specific staining

Question 11

Optimum temp for metallic stains

Answer 11

75–95°C

Question 12

Optimum temp for non-metallic stains

Answer 12

55–60°C

Question 13

Type of system

Answer 13

Enclosed, vacuum-assisted, batch mode

Question 14

Processing time in rapid processor

Answer 14

3 hours

Question 15

Tissue thickness requirement

Answer 15

1.5 mm

Question 16

Microwave wattage used

Answer 16

Low wattage

Question 17

Specimen capacity per hour

Answer 17

Up to 120 specimens

Question 18

8-hour shift cassette capacity

Answer 18

960 cassettes

Question 19

Time per 4-step cycle

Answer 19

15 minutes each per retort

Question 20

Is formalin used?

Answer 20

No – formalin- and xylene-free

Question 21

Quality of sections vs conventional

Answer 21

Comparable or superior

Question 22

IHC on rapid sections

Answer 22

Can be done without antigen retrieval

Question 23

Fixative for molecular testing

Answer 23

universal molecular fixative - UMFIX (methanol + PEG)

Question 24

UMFIX advantages

Answer 24

Non-volatile, room temp active, preserves DNA/RNA

Question 25

Tissue thickness for proper penetration

Answer 25

1.5 mm

Question 26

Special tissues requiring extra steps

Answer 26

Brain, large blocks

Question 27

Paraffin for microwave

Answer 27

Must be liquid – microwave can’t melt pellets

Question 28

Final step boiling point for alcohol

Answer 28

82°C for ethanol, 78°C for isopropanol

Question 29

Grossing time consideration

Answer 29

May increase due to thin sections

Question 30

Frozen section result time

Answer 30

15–30 minutes

Question 31

Frozen vs rapid processor result time

Answer 31

Frozen: faster (15–30 mins); Rapid: 4–5 hours

Question 32

Advantages of frozen section

Answer 32

Fast, minimal equipment, no formalin

Question 33

Disadvantage of frozen section

Answer 33

Poorer morphology than paraffin sections

Question 34

Frozen section uses

Answer 34

Tumor diagnosis, enzyme histochemistry, lipids, immunofluorescence, silver stains

Question 35

Most rapid freezing method

Answer 35

Liquid nitrogen

Question 36

Liquid nitrogen temp for cutting

Answer 36

–10°C to –25°C

Question 37

Liquid nitrogen disadvantages

Answer 37

Ice crystals, overcooling, uneven freezing

Question 38

Best for freezing muscle

Answer 38

Isopentane cooled by liquid nitrogen

Question 39

Other freezing methods

Answer 39

CO₂ gas, aerosol sprays

Question 40

Cold knife procedure uses

Answer 40

CO₂ gas

Question 41

Cold knife steps

Answer 41

Filter paper → tissue → freeze → cut at dew line

Question 42

Cold knife section removal

Answer 42

Camel hair brush or moistened finger

Question 43

Cryostat machine components

Answer 43

Tissue shelf, rotary microtome, blade, anti-roll plate

Question 44

Cryostat section behavior

Answer 44

Does not form ribbons

Question 45

Cryostat temperature

Answer 45

–18°C to –20°C

Question 46

Cryostat advantage

Answer 46

Same temp for knife, tissue, environment

Question 47

Mounting medium in Cryostat

Answer 47

O.C.T. compound

Question 48

Tissue freezing temps by type

Answer 48

-5°C to –35°C depending on tissue (e.g., brain, breast)

Question 49

Why freeze fixed tissue?

Answer 49

For fat/lipid or nerve special stains

Question 50

Slide coating for fixed frozen tissue

Answer 50

Albumin or chrome-glycerin jelly

Question 51

Fix before freezing for quick diagnosis

Answer 51

Boil in 10% buffered formalin for 1–2 minutes

Question 52

Special fixative for lipids

Answer 52

10% formol calcium at 4°C

Question 53

Wash alcohol-fixed tissue before freezing

Answer 53

12–24 hrs in water

Question 54

How biopsies are divided

Answer 54

1) Formalin-fixed (H&E), 2) Snap-frozen (cryostat), 3) Resin-embedded (EM/biochemical)

Question 55

Why use special processing?

Answer 55

Preserve enzyme activity and intact chemicals

Question 56

Most ideal for enzyme studies

Answer 56

Frozen section

Question 57

Other methods for special processing techniques

Answer 57

Freeze drying, freeze substitution

Question 58

Quenching

Answer 58

Rapid freezing using –160°C to –180°C (isopentane/propane)

Question 59

Desiccation and sublimation conditions

Answer 59

-30°C to –40°C in vacuum for 24–48 hrs

Question 60

Sublimation definition

Answer 60

Ice → gas (no liquid phase)

Question 61

Post-drying processing

Answer 61

Fix, embed, and stain

Question 62

Advantages of freeze-drying

Answer 62

Fresh tissue, minimal shrinkage, preserves enzymes

Question 63

Disadvantages of freeze-drying

Answer 63

Time-consuming, costly, brittle tissue

Question 64

Water to remove by weight

Answer 64

70–80%

Question 65

Used for studies like

Answer 65

Immunocytochemistry, hormones, fluorescent microscopy

Question 66

Freeze-substitution process

Answer 66

Use fixative + solvent instead of vacuum

Question 67

Freezing step

Answer 67

3:1 propane-isopentane at –175°C

Question 68

Cutting

Answer 68

8–10 µm in cryostat

Question 69

Transfer to

Answer 69

Water-free acetone at –70°C for 12h–1wk

Question 70

Advantages over freeze-drying

Answer 70

Cheaper and faster

Question 71

Good substituting fixatives

Answer 71

Osmium tetroxide, mercuric chloride, picric acid in alcohol