Basics of Immunohistochemistry

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Basics of Immunohistochemistry

• Vivien Schacht1 and Johannes S. Kern2
•  
• 1Department of Dermatology, Hannover Medical
  School, Hannover, Germany schacht.vivien_at_mh-hanno
  ver.de
• 2Department of Dermatology, Medical Center
  University of Freiburg, Freiburg, Germany
  johannes.steffen.kern_at_uniklinik-freiburg.de

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INTRODUCTION

• Immunohistochemistry (IHC)
• Demonstrates specific antigens in formalin-fixed,
  paraffin-embedded (FFPE) tissues.
• Antigen-antibody construct is visualized through
  light microscopy gt morphology of the tissue
  around the specific antigen is visible.
• Results are reported semiquantitatively
  (0) and have important diagnostic and
  prognostic implications.

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HISTORY

• 1940 immunofluorescence staining on frozen
  sections based on antigen-antibody interaction
  was presented by Coons.
• 1974 Taylor and Burns developed IHC on routinely
  processed FFPE tissues.
• 1975 Köhler and Milstein presented the hybridoma
  technique to produce monoclonal antibodies.
• gt broad range of antigens and high staining
  quality

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How Is Immunohistochemistry Performed?

• Tissue processing and epitope retrieval Formalin
  fixation Cutting sections Mount on glass
  slides Enzyme digestion
• Antigen-antibody interaction Direct or indirect
  method
• Visualization through different detection
  systems Fluorescent compounds or active enzymes

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Quality Control

• Purpose Sensitive and specific, reproducible
  and standardized
• Positive control Sample containing the antigen
  of interest
• Negative control Same sample as for the
  positive control, but primary antibody replaced
  by non-binding immunoglobulin
• Validation of IHC Round robin tests Staining
  various tissue and tumor types Comparing
  staining results of different antibodies that
  recognize similar proteins.

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Immunohistochemical Techniques
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Immunohistochemical Results for Select BRAF
V600E Mutation Positive Cases by DNA Analyses
Feller et al. 2013