Antigen Retrieval Using Sodium Hydroxide for Prion Immunohistochemistry in Bovine Spongiform Encephalopathy and Scrapie

Formalin-fixed and paraffin wax-embedded (FFPE) tissue sections are usually used for histopathological and immunohistochemical analyses in prion diseases in animals and man. However, formalin fixation cross-links proteins, reducing disease-associated prion protein (PrPSc) immunolabelling. To detect PrPSc in animals naturally affected with bovine spongiform encephalopathy (BSE) and scrapic, we applied minimal pretreatment with sodium hydroxide (NaOH). This simple pretreatment, combined with enzymatic digestion using proteinase K (PK), was equally effective in the detection of PrPSc in FFPE tissue, and superior in terms of speed, compared with the usual autoclaving method. The most effective results, without any section loss, were obtained with 10 mu g/ml PK in phosphate buffered saline containing 0.1% Triton-X at room temperature for 10 min and 150 niM NaOH at 60 degrees C for 10 min. By this simple procedure, PrPSc was visualized in the brain of animals with BSE and scrapie using a range of anti-PrP primary antibodies. (C) 2010 Elsevier Ltd. All rights reserved.