1. Basics of Immunohistochemistry
Vivien Schacht1 and Johannes S. Kern2
1Department of Dermatology, Hannover Medical School, Hannover, Germany;
2Department of Dermatology, Medical Center – University of Freiburg, Freiburg, Germany;

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

3. 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.
=> broad range of antigens and high staining quality

4. 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

5. 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.

6. Immunohistochemical Techniques
Direct method
Indirect method
with polymer chain detection system

7. in Human Melanocytic Tumors
p16INK4a Expression
in Human Melanocytic Tumors
Scurr et al. 2011

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