title: Heat-induced Epitope Retrieval for Immunohistochemistry at High Altitude: A Quality Assurance Study.
abstract: Heat-induced epitope retrieval (HIER) of formalin-fixed paraffin-embedded tissues is now a standard practice in immunohistochemistry (IHC). In this study, we aimed to test the effect of altering HIER temperature on IHC staining quality at high altitude, the hypothesis being that lower HIER temperatures would result in improved staining patterns.
  
  
  
  In a laboratory at high altitude (Aurora, CO), we used a platform with automated onboard epitope retrieval, and systematically tested 3 different HIER temperatures (100°C, 95°C, 90°C) with 4 IHC stains that are commonly used in routine practice: CD3, Ki67, CK20, and Melan A (n=10 for each antibody/epitope retrieval temperature combination). A scoring system was devised, the slides were scored in a blinded manner, and statistical analysis was performed. For comparison, the same study was performed in a laboratory near sea level (Atlanta, GA).
  
  
  
  At high altitude, lower HIER temperatures resulted in improved staining patterns, as quantified by stronger staining intensity and greater area of the slides stained. The scores obtained with HIER temperatures of 95°C and 90°C were higher than those obtained with HIER of 100°C, and the difference was found to be statistically significantly for some antibody/epitope retrieval temperature combinations (P<0.05). This effect was not seen in the laboratory near sea level.
  
  
  
  We show that alternate epitope retrieval recommendations are warranted for laboratories at high altitude. Furthermore, we suggest that manufactures should consider how their instruments will perform at high altitude as they further automate the process of IHC.
authors: *Department of Pathology and Laboratory Medicine, Emory University School of Medicine, Atlanta, GA †Arizona State University, Phoenix, AZ ‡Ruegg IHC Consulting, Bennett, Aurora, CO.
authors: Keith A
authors: KA
authors: Stevens
authors: Charles K
authors: CK
authors: Kovach
authors: Michael
authors: M
authors: Tompkins
authors: Patsy
authors: P
authors: Ruegg
authors: Cynthia
authors: C
authors: Cohen
mesh_terms: D000531:Altitude; D017252:CD3 Complex / Q000737:chemistry; D006358:Hot Temperature; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods* / Q000592:standards; D053554:Keratin-20 / Q000737:chemistry; D019394:Ki-67 Antigen / Q000737:chemistry; D058965:MART-1 Antigen / Q000737:chemistry; D016612:Paraffin Embedding; D011786:Quality Control
chemical_list: D017252:CD3 Complex; C507499:KRT20 protein, human; D053554:Keratin-20; D019394:Ki-67 Antigen; D058965:MART-1 Antigen