title: Quantification of human diamine oxidase.
abstract: OBJECTIVE
  Diamine oxidase (DAO) is essential for extracellular degradation of histamine. For decades activity assays with inherent limitations were used to quantify the relative amounts of DAO. No reference DAO standard is available. Absolute DAO amounts cannot be determined. Controversy exists, whether DAO is circulating or not in non-pregnant individuals. The role of DAO as biomarker in various diseases is ambiguous. It is not clear, whether precise quantification of human DAO antigen using commercially available enzyme-linked immunosorbent assays (ELISAs) is possible. The objective was to develop a precise and robust ELISA to quantify DAO in various biological fluids.
  
  
  METHODS
  A research prototype ELISA was established using a mouse monoclonal antibody for capturing and a polyclonal rabbit serum IgG fraction for the detection of human DAO. The limit of blank (LoB), limit of detection (LoD) and estimated limit of quantification (eLoQ) and normal DAO concentrations in serum and plasma were determined.
  
  
  RESULTS
  The LoB, LoD and eLoQ derived from 42 standard curves are 0.27, 0.48 and 0.7ng/mL respectively. The detection range using the LoD as the lower and the highest DAO standard as the upper boundary is 0.5 to 450ng/mL. Serum and plasma mean/median concentrations are between 0.5 and 1.5ng/mL in healthy volunteers (n=58) and mastocytosis patients (n=19) and plateau at approximately 145ng/mL (n=16) during pregnancy. Accurate quantification was not influenced by heparin (DAO is a heparin-binding protein), lipaemic or hemolytic serum. The measured DAO antigen concentrations are in close agreement with published enzymatic activity data using radioactive putrescine as substrate.
  
  
  CONCLUSIONS
  This research prototype ELISA is able to reliably and accurately quantify human DAO in different biological fluids. The potential of DAO as biomarker in various diseases can be evaluated.
authors.affiliation: Department of Clinical Pharmacology, Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austria. Electronic address: thomas.boehm@meduniwien.ac.at.
authors.forename: Thomas
authors.initials: T
authors.lastname: Boehm
authors.affiliation: Department of Obstetrics and Gynecology, Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austria.
authors.forename: Sophie
authors.initials: S
authors.lastname: Pils
authors.affiliation: Department of Clinical Pharmacology, Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austria; Department of Biotechnology, University of Natural Resources and Life Sciences, Muthgasse 18, 1190 Vienna, Austria.
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Gludovacz
authors.affiliation: Department of Clinical Pharmacology, Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austria.
authors.forename: Helen
authors.initials: H
authors.lastname: Szoelloesi
authors.affiliation: Department of Clinical Pharmacology, Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austria.
authors.forename: Karin
authors.initials: K
authors.lastname: Petroczi
authors.affiliation: Institute of Immunology, Center for Pathophysiology, Infectiology and Immunology Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austria.
authors.forename: Otto
authors.initials: O
authors.lastname: Majdic
authors.affiliation: Department of Biomedical Sciences, Veterinary Research Tower, Cornell University, Ithaca, NY 14853-6401, USA.
authors.forename: Andrea
authors.initials: A
authors.lastname: Quaroni
authors.affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences, Muthgasse 18, 1190 Vienna, Austria.
authors.forename: Nicole
authors.initials: N
authors.lastname: Borth
authors.affiliation: Department of Internal Medicine I, Division of Hematology, Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austria.
authors.forename: Peter
authors.initials: P
authors.lastname: Valent
authors.affiliation: Department of Clinical Pharmacology, Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austria.
authors.forename: Bernd
authors.initials: B
authors.lastname: Jilma
mesh_terms: D006631:Amine Oxidase (Copper-Containing) / Q000097:blood*; D000818:Animals; D058846:Antibodies, Monoclonal, Murine-Derived / Q000737:chemistry; D015415:Biomarkers / Q000097:blood; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005260:Female; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry; D008297:Male; D051379:Mice; D011247:Pregnancy; D011817:Rabbits
chemical_list: D058846:Antibodies, Monoclonal, Murine-Derived; D015415:Biomarkers; D007074:Immunoglobulin G; D006631:Amine Oxidase (Copper-Containing)
keywords: Anaphylaxis; Diamine oxidase; Diamine oxidase quantification; Histamine degradation; Mastocytosis; Prognostic marker