350 - Topic 4 (Frozen Sections) Flashcards
how long does it take to produce a diagnostic-quality slide in frozen sectioning?
20 mins (CAP accreditation standard time)
Four major applications for frozen sections
- intraoperative consultations
- enzyme histochemistry
- immunofluorescence
- lipid stains
describe: intraoperative consultation
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
describe: enzyme histochemistry
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
describe: immunofluorescent techniques
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
describe: lipid stains
tissue processing uses solvents that dissolve lipids
frozen sections must be used to visualize lipids in tissue sections
what happens if tissue is frozen slowly?
large crystals form
esp. true for striated muscle
what is freezing artifact?
LARGE CRYSTALS FROM FREEZING SLOWLY THAT DISTORT tissue morphology and leave large holes in the section after staining
how is tissue freezing accomplished?
- heat extractors (metal plates and hammers found in cryostat)
- dry ice or container of isopentane (2-methylbutane) cooled with liquid nitrogen
- fast freezing!
temp of cryostat
-20C
cryostat temp for dense, highly cellular tissues
warmer temps = -10 to -15C
cryostat temp for open and connective tissues (esp. adipose)
cooler temps = -25 to -35C
T or F. Lower temps result in firmer blocks
T
frozen sectioning compound
FSC
- TISSUE ORIENTED IN THIS
- A VISCOUS LIQUID THAT QUICKLY FREEZES TO SUPPORT TJE TSSU AND FACILITATE SECTIONING
FSC analogous to what in microtomy
paraffin used to embed tissue
what happens after frozen sectioning?
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)
what happens once slides have been examined by a pathologist?
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
why do we compare paraffin-embedded tissues to prior frozen sections?
quality assurance!
frozen sections cannot and should not be used for final diagnosis due to their inferior quality
appropriate PPE for frozen sectioning
N95
goggles
gloves
gown
potentially infectious as tissue has not been fixed!!!
chatter
regular pattern of horizontal lines in tissue section (venetian blind)
due to tissue or block being too cold
OR loose component of cryotome
shattering
AKA fragmenting
may occur if cryostat temp too cold for tissue
common when sectioning lymph nodes
compression
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
lines/scores
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
how to correct static electricity in cryostat?
humidify the room or wipe down interior of cryotome with alcohol
cryostat should be disinfected at the end of the day using this
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)
this helps avoid buildup of ice crystals
automatic defrost cycle
most common cause of overheating crystats
dusty fins/coils
-> periodic vacuuming or dusting
