Application of xanthine oxidase-catalyzed luminol chemiluminescence in a mouse interleukin-5 immunoassay

A chemiluminescent substrate reagent for use in a sandwich immunoassay for the model antigen mouse interleukin-5 (IL-5) was developed using xanthine oxidase and luminol. Various parameters involved in this chemiluminescent reaction have been studied, including the substrate hypoxanthine, luminol and...

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Bibliographic Details
Published in:Journal of immunological methods 1996-10, Vol.197 (1), p.161-169
Main Authors: Rongen, H.A.H., van der Horst, H.M., van Oosterhout, A.J.M., Bult, A., van Bennekom, W.P.
Format: Article
Language:English
Subjects:
Animals
Biological and medical sciences
Edetic Acid - pharmacology
Enzyme-Linked Immunosorbent Assay - methods
Ferrous Compounds - pharmacology
Fundamental and applied biological sciences. Psychology
Fundamental immunology
Horseradish Peroxidase
IL-5
Immunoassay
Interleukin-5 - analysis
Luminescent Measurements
Luminol
Mice
Molecular immunology
Techniques
Xanthine Oxidase
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Summary:A chemiluminescent substrate reagent for use in a sandwich immunoassay for the model antigen mouse interleukin-5 (IL-5) was developed using xanthine oxidase and luminol. Various parameters involved in this chemiluminescent reaction have been studied, including the substrate hypoxanthine, luminol and the Fe(II)-EDTA complex. Addition of the Fe(II)-EDTA complex enhances the chemiluminescence signal considerably. The xanthine oxidase-catalyzed chemiluminescent immunoassay was compared to horseradish peroxidase-linked immunoassays with luminol as chemiluminescent, and tetramethyl benzidine as colorimetric substrate. The detection limit of the xanthine oxidase-luminol assay was found to be about 0.6 pg/ml IL-5, whereas the peroxidase-catalyzed immunoassays have detection limits of about 1.3 (HRP-TMB) and 2.9 pg/ml (HRP-luminol) IL-5.
ISSN:0022-1759
1872-7905
DOI:10.1016/0022-1759(96)00131-7