Introduction. Basic histological techniques. Histological staining (acid, basic, histochemistry). Všeobecné lekárstvo MUDr. Pavol Janega MUDr. Svetoslav Štvrtina MUDr. Peter Michalka Lucia Donárová Ústav patologickej anatómie LFUK a UN pracovisko Staré mesto Sasinkova 4, Bratislava Prof. MUDr. Ľudovít Danihel, CSc.
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• perserve E activity during fixation and processing
• frozen sections
Tissue with active enyzme substrate
Intermediate product
percipitate
reactant
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• The using of immunological methods in addition to morphological evaluation in the diagnosis of diseases allows the demonstration of various proteins production in tissues
• Evaluation of the presence, localization and quantification of specific proteins (Ag) by a specific antibody (Ab)
• Significance
• The diagnosis (presence or absence of specific markers)
• The prognosis (prognostic markers)
• The therapy (proof of markers having a relationship to the effectiveness of treatment, affecting resistance, adequacy of the used treatment)
Method
• Reliable and reproducible
• Low costs
Currently, the gold standard of diagnosis
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primary antibody
secondary antibody (anti-rabbit, mouse, goat,...)
antigen
Fluorescent stain
fluorescence method
positive = flourescens in fluorescent microscope
primary antibody (rabbit, mouse, goat,...)
secondary antibody (anti-rabbit, mouse, goat,...)
antigen
Enzyme (peroxidase)
Dye (DAB) precipitate (brown color) in light microscope
enzymatic method
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• Diagnostic markers (cytokeratins, vimentin...)
• Prognostic markers (Ki67...)
• Therapeutic markers (Her2...)
CYTOKERATINS
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• Epithelial cells
VIMENTIN
• Mezenchymal cells
AE1/3 cytokeratíny
KI67
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• Positive in proliferating cells in late G1,S,G2 and M phase, but not in G0 and early G1