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Biochemical Measurement of Neonatal Hypoxia
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Quantification of human diamine oxidase.

Thomas Boehm1, Sophie Pils2, Elisabeth Gludovacz3

  • 1Department of Clinical Pharmacology, Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austria.

Clinical Biochemistry
|January 3, 2017
PubMed
Summary

A new enzyme-linked immunosorbent assay (ELISA) precisely quantifies diamine oxidase (DAO) levels in biological fluids. This assay overcomes previous limitations, enabling accurate DAO measurement for biomarker research in various diseases.

Keywords:
AnaphylaxisDiamine oxidaseDiamine oxidase quantificationHistamine degradationMastocytosisPrognostic marker

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Area of Science:

  • Biochemistry
  • Immunology
  • Clinical Chemistry

Background:

  • Diamine oxidase (DAO) is crucial for histamine degradation, but its quantification has been limited by activity assays lacking standards.
  • Previous methods could not determine absolute DAO amounts, leading to ambiguity regarding its presence in non-pregnant individuals and its role as a disease biomarker.
  • The precise quantification of human DAO antigen using enzyme-linked immunosorbent assays (ELISAs) remained unclear.

Purpose of the Study:

  • To develop a precise and robust ELISA for quantifying human diamine oxidase (DAO) antigen in various biological fluids.
  • To establish the analytical performance characteristics of the developed ELISA, including limits of detection and quantification.
  • To determine normal DAO concentrations in serum and plasma and assess its levels during pregnancy.

Main Methods:

  • A research prototype ELISA was developed using a mouse monoclonal antibody for capture and a rabbit polyclonal antibody IgG fraction for detection.
  • Analytical sensitivity was assessed by determining the limit of blank (LoB), limit of detection (LoD), and estimated limit of quantification (eLoQ).
  • DAO concentrations were measured in serum and plasma from healthy volunteers, mastocytosis patients, and pregnant individuals.

Main Results:

  • The developed ELISA demonstrated robust analytical sensitivity with LoB, LoD, and eLoQ of 0.27, 0.48, and 0.7 ng/mL, respectively.
  • The assay accurately quantified DAO in serum and plasma, with normal concentrations ranging from 0.5 to 1.5 ng/mL in healthy individuals and patients.
  • DAO levels significantly increased during pregnancy, plateauing around 145 ng/mL, and quantification was unaffected by heparin, lipemia, or hemolysis.

Conclusions:

  • The developed research prototype ELISA reliably and accurately quantifies human DAO antigen in diverse biological fluids.
  • This assay provides a valuable tool for overcoming previous limitations in DAO measurement.
  • The validated ELISA enables the accurate evaluation of DAO's potential as a biomarker in various physiological and pathological conditions.