title: Combining immunodetection with histochemical techniques: the effect of heat-induced antigen retrieval on picro-Sirius red staining.
abstract: Picro-Sirius red is a routine diagnostic stain intended for the histological visualization of collagen fibers (fibrosis) in tissue. Multi-label immunohistochemistry is a powerful tool used by researchers to visualize different cell types and their location within a tissue specimen, and to observe co-localization of antigens. Combining the specificity of immunodetection with the simplicity of Sirius red staining will allow researchers to visualize multi-antigen detection in relation to fibrosis, a common histological feature of injury in many chronic diseases. Pre-treatment of formalin-fixed, paraffin-embedded tissue (FFPE) specimens with antigen retrieval is essential for the work-up of most commercially available antibodies. The most common form of antigen retrieval involves boiling tissue specimens in buffer to break the cross-linkages caused by formalin fixation. However, this method causes tissue modification and collagen fiber shrinkage leading to suboptimal results when counterstaining for Sirius red. Reduced heat and enzymatic digestion are antigen retrieval methods compatible with Sirius red counterstaining. This paper will discuss the difficulties faced when combining these two staining methods, and provide a detailed method for the simultaneous detection of antigen and Sirius red in FFPE tissues.
authors.affiliation: Centre for Liver Disease Research, School of Medicine, The University of Queensland, Translational Research Institute, Brisbane, Australia (VLG) v.gadd@uq.edu.au.
authors.forename: Victoria L
authors.initials: VL
authors.lastname: Gadd
mesh_terms: D000941:Antigens / Q000032:analysis*; D001391:Azo Compounds / Q000032:analysis; D003094:Collagen / Q000032:analysis; D004396:Coloring Agents / Q000032:analysis; D005404:Fixatives; D005557:Formaldehyde; D006358:Hot Temperature; D007150:Immunohistochemistry / Q000379:methods*; D016612:Paraffin Embedding; D013194:Staining and Labeling / Q000379:methods*; D016707:Tissue Fixation
chemical_list: D000941:Antigens; D001391:Azo Compounds; D004396:Coloring Agents; D005404:Fixatives; C009798:C.I. direct red 80; D005557:Formaldehyde; D003094:Collagen
keywords: HIAR; Immunohistochemistry; Sirius red; antigen retrieval; histology