Background staining of visualization systems in immunohistochemistry: comparison of the Avidin-Biotin Complex system and the EnVision+ system

Appl Immunohistochem Mol Morphol. 2007 Mar;15(1):103-7. doi: 10.1097/01.pai.0000213102.33816.13.

Abstract

The aim of this study was to evaluate specific immunostaining and background staining in formalin-fixed, paraffin-embedded human tissues with the 2 most frequently used immunohistochemical detection systems, Avidin-Biotin-Peroxidase (ABC) and EnVision+. A series of fixed tissues, including breast, colon, kidney, larynx, liver, lung, ovary, pancreas, prostate, stomach, and tonsil, was used in the study. Three monoclonal antibodies, 1 against a nuclear antigen (Ki-67), 1 against a cytoplasmic antigen (cytokeratin), and 1 against a cytoplasmic and membrane-associated antigen and a polyclonal antibody against a nuclear and cytoplasmic antigen (S-100) were selected for these studies. When the ABC system was applied, immunostaining was performed with and without blocking of endogenous avidin-binding activity. The intensity of specific immunostaining and the percentage of stained cells were comparable for the 2 detection systems. The use of ABC caused widespread cytoplasmic and rare nuclear background staining in a variety of normal and tumor cells. A very strong background staining was observed in colon, gastric mucosa, liver, and kidney. Blocking avidin-binding capacity reduced background staining, but complete blocking was difficult to attain. With the EnVision+ system no background staining occurred. Given the efficiency of the detection, equal for both systems or higher with EnVision+, and the significant background problem with ABC, we advocate the routine use of the EnVision+ system.

Publication types

  • Comparative Study
  • Evaluation Study

MeSH terms

  • Antibodies, Monoclonal
  • Avidin
  • Biotin
  • Humans
  • Immunohistochemistry / methods*
  • Immunohistochemistry / standards
  • Peroxidase
  • Reagent Kits, Diagnostic / standards*
  • Staining and Labeling
  • Tissue Fixation

Substances

  • Antibodies, Monoclonal
  • Reagent Kits, Diagnostic
  • Avidin
  • Biotin
  • Peroxidase