Transcript Update on Immunohistochemistry 2014

Cell block:
Monokeratin – rare dot
On Slide Control
Tissue biopsy:
Monokeratin – positive
Changing the type of fixation
Pre-Analytical (surgery → unstained section)
results in downstream
•Ischemic time
problems in the analytical and
•Type of fixation
post-analytical phases !
•Length of fixation
Analytical (unstained section → coverslip)
•Decalcification
•Antibody supplier
•Processing
•Clone
•Dehydration and clearing
•Epitope retrieval
•Paraffin impregnation
•Dilution
•Paraffin sectioning
•Amplification
•Storage
•Chromogen
Post-Analytical (interpretation)
•Counterstain
•Design of controls
•Detection
•Positive controls (internal and external)
•Blocker
•Negative controls
•Mounting
•Interpretation
•Critical indicators
•Reporting
This implies that a laboratory must have supply
of control tissue developed for each of these
situations with IHC protocols developed for each
antibody on these tissues
Let’s be honest – who actually does this?
Am J Clin Pathol 2010;133:354-365
Immunohistochemical Practices of Cytopathology Laboratories: A
survey of Participants in the College of American Pathologists
Nongynecologic Cytopathology Education Program.
Arch. J. Clin. Pathol. 2010; 133: 354-365
Arch. J. Clin. Pathol. 2010; 133: 354-365
Arch. J. Clin. Pathol. 2010; 133: 354-365
TTF1 Expression in Normal Lung Tissue
CytoLyt® fixation and decalcification pretreatments alter antigenicity in normal tissues compared
to standard formalin fixation. Jennette R. Gruchy, Penny J. Barnes, Kelly A. Dakin Hache.
Applied Immunohistochemistry and Molecular Morphology, 2014, in press.
TTF1 FFPE
TTF1 CytoLyte™
CD20 Expression in Normal Appendix
CytoLyt® fixation and decalcification pretreatments alter antigenicity in normal tissues compared
to standard formalin fixation. Jennette R. Gruchy, Penny J. Barnes, Kelly A. Dakin Hache.
Applied Immunohistochemistry and Molecular Morphology, 2014, in press.
CD20 FFPE
CD20 CytoLyte™
D2-40 Expression in Normal Appendix
CytoLyt® fixation and decalcification pretreatments alter antigenicity in normal tissues compared
to standard formalin fixation. Jennette R. Gruchy, Penny J. Barnes, Kelly A. Dakin Hache.
Applied Immunohistochemistry and Molecular Morphology, 2014, in press.
D2-40 FFPE
D2-40 CytoLyte™
Cell block:
Monokeratin – rare dot
Tissue biopsy:
Monokeratin – positive
Monokeratin control appropriate on both
cytology cell block slide and tissue biopsy
slide
Cell block:
TTF-1 – negative
Tissue biopsy:
TTF-1 – positive
TTF-1 control appropriate on both cytology
cell block slide and tissue biopsy slide
Cell block:
Synaptophysin – focal weak
Tissue biopsy:
Synaptophysin – positive
Synaptophysin control appropriate on both
cytology cell block slide and tissue biopsy
slide
DX: small cell carcinoma
• On-slide tissue control shows that the slide receives the antibody of
interest
• On-slide tissue control cannot detect false negative reaction if control
tissue is not fixed and processed in the same manner as the patient
tissue
• Placing formalin fixed control tissue on the slide gives a false sense of
security
• Formalin fixed controls are not valid for cytology specimens