Frozen sections and other intraoperative

Intraoperative histopathology consultations

1. Frozen Sections and Other Intraoperative
Consultations

2. INTRODUCTION
Intraoperativeconsultations fall into two general categories.
•Microscopic consultations: -Usually performed as frozen
sections.
-Can also be performed using
touch preparations.
•Nonmicroscopicconsultations are gross examinations of a
specimen that provide the surgeon with real-time information
on tissue margins and the anatomic extent of disease
processes.

3. What is frozen section?
•The frozen section procedure is a pathological laboratory
procedure to perform rapid microscopic analysis of a
specimen.
•The technical name for this procedure is Cryosection.

4. History:
•Frozen Section Resulted
from need for Rapid Intra
operative diagnosis.
•Mayo Clinic Chief of
Pathology Louis B. Wilson
pioneered the frozen section
at the Rochester,Minnesota
clinic in 1905 .

6. Section of the tissue:
1) Tissue sample should represent the specimen.
2) Should not contain any necrotic area.
3) 3-4mm is ideal section.
4) Sample once collected should be frozen immediately.

7. The ideal temperature for different tissues

9. FIXATION
• For fast result, it requires to cut sections of frozen but
unfixed tissue.
• ‘Short period of fixations before freezing makes section
cutting easier and yields better sections’ (especially for
fat/mucincontaining tissue)
• Ideal fixative is 10% formal-saline for 10 minsat 60°C

10. Freezing techniques:
• Liquid nitrogen (-1900c)
• Isopentanecooled by liquid nitrogen (-1500c)
• Dry ice(-700c)
• Carbon dioxide gas(-700c)
• Aerosol sprays(-500c)

11. Methods for suitable freezing
1)Cryostat:
-microtome is inside the chamber
-microtome in under constant temperature control
2)Freezing microtome:
-Tissue fixed separately
-Microtome is not under temperature control

12. Cryostat Machine

13. Cryostat: Types:
1)Closed-cycle cryostats
2)continuous-flow cryostat
3)Bath cryostat
4)Multistage cryostats

14. Frozen section embedding medium

15. Taking of sections from stage
•The cut section will rest on the surface of the blade holder.
•After cutting ,a room temperature slide is held above the
section.
•The electrostatic attraction causes the tissue to adhere to the
slide.

16. Staining:
1)Rapid H-E method
2)Toludinemethod
3)MethyleneBlue
4)Methyl violet for amyloid
5)Oil red O for fat
6)PAS

17. Technique of staining
• Immediately fix frozen section in 95% ethyl alchohol
• Rinse well in distilled water
• Stain with HematoxylinStain
• Wash well in distilled water
• Counterstainin Eosin Y Working Solution
• Dehydrate in ethyl alcohol
• Subsequent treatment

18. Procedure

19. Pros and cons of frozen section
Pros:
• Fastest of all methods
• Excellent for IHC, IF, ISH.
• Often easiest to section-depending upon the tissue.
Cons:
• Poorest morphology
• Prone to freezing artifact-must be snap frozen

20. Common problems: artifacts
• Ice crystal artifacts:
-Cause: slow freezing of tissue
-Solution:Freeze
fast(flash/snap)
-It is a chemical property of
water, that water will expand
on freezing.

21. Common problems: artifacts

22. Common problems: artifacts
•Over freezing can cause
sections to have holes.
•Tissues with high water
content eg. edematous or
bloody tissue will show this
Problem.

23. Quality assurance
• Never two cases in one grossing area
• System of identification of block and slide should be established
• Never two cases in one cryostat
• Unlabeled slides should never be used
•Correlation has to be done in each case.
• The cryostat must be clean periodically.
• Less frequently used machine can be decontaminated on a longer
cycle
• Records keeping.

24. Frozen sections:
Indications:
❑To establish a tissue diagnosis (e.g., to confirm the presence of
parathyroid tissue in a parathyroidectomyspecimen).
❑Determine the nature of a lesion that may require ancillary
testing that requires special fixatives or media.
(e.g., RPMI for flow cytometryor glutaraldehydefor electron
microscopy).

25. Frozen sections:
Indications:
❑Establish that sufficient diagnostic tissue has been obtained,
❑Identify metastatic disease,
❑Assess surgical margins or extent of disease.

26. Approximate time taken
•In optimum working conditions and experienced hands, the
entire consultation can often be performed in 10 to 15 minutes
from the time of the arrival of the specimen in the frozen
section room to the notification of the surgeon of the
diagnosis.

27. The frozen section procedure:
•Frozen sections are performed by freezing the tissue in a block
of specialized embedding medium.
•Followed by cutting thin (usually 5 mm) sections from the
block using a cryostat (refrigerated microtome).
•The sections are adhered to glass slides, fixed in ethanol, and
stained with hematoxylinand eosin (H&E).

28. The frozen section procedure:
•Small specimens may be completely utilized for frozen
section slide preparation, but if possible, a portion of the tissue
should be preserved for routine handling to avoid freezing
artifacts that can compromise interpretation of the permanent
sections (e.g., in the case of brain biopsies).
•For larger tissue samples, judgment must be exercised in
gross sampling so that the area(s) of highest diagnostic yield is
selected for frozen section.
•Cytological imprints from tissues can be an important adjunct
in diagnosis, especially for hematolymphoidabnormalities,
lymph node biopsies, and thyroid lesions.

29. Interpretation
The interpretation of frozen sections requires integration of
the:
•Histologicmorphology in the H&E-stained sections;
•The gross features of the specimen;
•Information from the surgeon regarding the origin of the
tissue,
•Indication for the consultation (including the clinical history,
radiological findings, and intraoperativeobservations);
•The ways in which the frozen section diagnosis will affect the
operative strategy.

30. Interpretation
Note:
❑In some difficult cases, it may be necessary to request
additional tissue for frozen section analysis.
❑If additional tissue cannot be obtained, deferral of a definitive
diagnosis pending examination of formalin-fixed paraffin-
embedded sections is acceptable.

31. Communication of findings
•Clear communication of a concise diagnosis to the surgeon
is the last step of the intraoperativeconsultation.

32. Accuracy of frozen sections
•The accuracy of frozen sections will vary from institution
to institution on the basis of the types of surgical cases
evaluated and the experience of the involved pathologists.
•Regular self-audits of the frozen section service are desirable
so that surgeons and pathologists are aware of the performance
characteristics of the modality in their own hands.