350 - Topic 4 (Frozen Sections)

Question 1

how long does it take to produce a diagnostic-quality slide in frozen sectioning?

Answer 1

20 mins (CAP accreditation standard time)

Question 2

Four major applications for frozen sections

Answer 2

• intraoperative consultations
• enzyme histochemistry
• immunofluorescence
• lipid stains

Question 3

describe: intraoperative consultation

Answer 3

before closing a patient during surgery, the doctor may ask histology to identify an unexpected finding or confirm t hat a biopsy is negative for cancer

Question 4

describe: enzyme histochemistry

Answer 4

enzymes degrade rapidly when removed from blood supply and enzyme activity is greatly reduced after chemical fixation
- freezing tissue = best option for preservation and detection of enzymes
- esp. true for muscle biopsies

Question 5

describe: immunofluorescent techniques

Answer 5

antibodies with fluorescent labels are used to detect labile antigens
- chem. fixation may alter or denature these agents = no longer react with Ab
- aldehyde fixative may also create autofluorescence

Question 6

describe: lipid stains

Answer 6

tissue processing uses solvents that dissolve lipids
frozen sections must be used to visualize lipids in tissue sections

Question 7

what happens if tissue is frozen slowly?

Answer 7

large crystals form
esp. true for striated muscle

Question 8

what is freezing artifact?

Answer 8

LARGE CRYSTALS FROM FREEZING SLOWLY THAT DISTORT tissue morphology and leave large holes in the section after staining

Question 9

how is tissue freezing accomplished?

Answer 9

• heat extractors (metal plates and hammers found in cryostat)
• dry ice or container of isopentane (2-methylbutane) cooled with liquid nitrogen
• fast freezing!

Question 10

temp of cryostat

Answer 10

-20C

Question 11

cryostat temp for dense, highly cellular tissues

Answer 11

warmer temps = -10 to -15C

Question 12

cryostat temp for open and connective tissues (esp. adipose)

Answer 12

cooler temps = -25 to -35C

Question 13

T or F. Lower temps result in firmer blocks

Answer 13

T

Question 14

frozen sectioning compound

Answer 14

FSC
- TISSUE ORIENTED IN THIS
- A VISCOUS LIQUID THAT QUICKLY FREEZES TO SUPPORT TJE TSSU AND FACILITATE SECTIONING

Question 15

FSC analogous to what in microtomy

Answer 15

paraffin used to embed tissue

Question 16

what happens after frozen sectioning?

Answer 16

fixed briefly in formalin or alcohol (formalin»)
- can be left to dry for future staining or rinsed in water for staining solutions
- intraoperative consult = slides stained with H&E or Dif-Quik (commercial Romanowsky stain)

Question 17

what happens once slides have been examined by a pathologist?

Answer 17

if no further sections are required, the tissue is thawed, rinsed wit water to remove FSC and fixed in formalin prior to routine overnight processing
- next day = slide is cut from parafin block to compare to frozen sections

Question 18

why do we compare paraffin-embedded tissues to prior frozen sections?

Answer 18

quality assurance!
frozen sections cannot and should not be used for final diagnosis due to their inferior quality

Question 19

appropriate PPE for frozen sectioning

Answer 19

N95
goggles
gloves
gown
potentially infectious as tissue has not been fixed!!!

Question 20

chatter

Answer 20

regular pattern of horizontal lines in tissue section (venetian blind)
due to tissue or block being too cold
OR loose component of cryotome

Question 21

shattering

Answer 21

AKA fragmenting
may occur if cryostat temp too cold for tissue
common when sectioning lymph nodes

Question 22

compression

Answer 22

when smaller than the face of the block
- have a tendency to collapse onto edge of blade when cutting
- block too warm OR blade too dull

Question 23

lines/scores

Answer 23

artifact noted macroscopically
vertical lines (perpendicular to blade) noted
- defect in blade probs
- calcification in tissue may also be culprit
- try moving to new area of blade, if persists = defect in block

Question 24

how to correct static electricity in cryostat?

Answer 24

humidify the room or wipe down interior of cryotome with alcohol

Question 25

cryostat should be disinfected at the end of the day using this

Answer 25

70% ethanol
- but not high-level disinfectant so cryostat should be defrosted and decontaminated with more powerful = Oxivir or CaviCide
- older cryostat = heating chamber to vaporize glutaraldehyde or formalin
or UV light (cannot penetrate debris though)

Question 26

this helps avoid buildup of ice crystals

Answer 26

automatic defrost cycle

Question 27

most common cause of overheating crystats

Answer 27

dusty fins/coils
-> periodic vacuuming or dusting