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Frozen section

Troubleshooting artifacts and poor technique


Author: Jessica Wallace (see Reviewers page)
Revised: 23 October 2011, last major update October 2011
Copyright: (c) 2011, PathologyOutlines.com, Inc.

Ice crystal artifacts
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● Due to slow freezing of tissue
Solution: Freeze fast (flash/snap); the faster the freeze, the smaller the ice crystals, the less tissue damage (best freezing method is arguably liquid nitrogen)
● Smaller tissues yield less artifact - optimally tissue should be 0.5 x 0.5 x 0.3 cm or less
● Never freeze fragments larger than the diameter of the chuck
● Avoid freezing fat around tissue
● Blot the outer surface of the tissue dry with gauze before making your block

   
Left: ice crystals in edematous stroma by frozen section, right: H&E

   
Nuclear ice crystals (particularly a problem with thinner sections): left-lung adenocarcinoma, right-uterine sarcoma

       
Glioma

Knife artifact
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● A nicked cutting blade will produce a split/tear in your section
Solution:: change your blade every few cases; some institutions use a new blade for each case


Breast tissue

Over / underfreezing
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● Overfreezing can cause section to have holes
● Solution: polish block with a couple extra turns of the blade to create friction and warm up block by pressing on it with your finger (5-10 seconds)
● Underfreezing can be troublesome for fatty tissue
Solution: add heat sink to block or select rapid freeze setting on your cryostat (if available)

Staining issues
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● Dirty "stain line" can cause floaters (extraneous foreign tissue) to adhere to slides; overly diluted stains and alcohols can diminish slide quality
● Poor staining hinders frozen section diagnoses, as nuclear detail is compromised
Solutions: (a) maintain a clean stain line by frequent solution changes; (b) follow recommended staining times; (c) don't rush
● Note: brain tissue may stain best in eosin for 60+ seconds
● Water: should be changed after each frozen section
● Alcohols and stains: change at least weekly, alcohols may need to be changed more frequently depending on work load

Fatty tissue
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● Includes lymph nodes, breast, skin; may be too soft to cut
Solution: maintain an extremely cold cutting temperature (-20C)
● Firm lymph nodes, spleen, brain and liver cut better at -10C; tissue may shatter if sectioning is performed at lower temps

Air bubbles
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● May be trapped under cover slips, which can cause the underlying tissue to dry out
Solution: make sure an appropriate amount of resin (2 drops) is applied; gently move air bubbles off the slide with finger or tweezers; do not press on the slide too hard or it will break

Overly thick sections
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● May cause tissue to fall off slide
Solution: reduce the cryostat's sectioning thickness

End of Frozen section > Troubleshooting artifacts and poor technique


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