Journal of Biotechnology 2012-11-15

An automated multiplex specific IgE assay system using a photoimmobilized microarray.

Yoshihiro Ito, Nozomi Moritsugu, Takahisa Matsue, Kiyomi Mitsukoshi, Hirohito Ayame, Norihiko Okochi, Hideshi Hattori, Hideo Tashiro, Sakura Sato, Motohiro Ebisawa

Index: J. Biotechnol. 161(4) , 414-21, (2012)

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Abstract

An automated microarray diagnostic system for specific IgE using photoimmobilized allergen has been developed. Photoimmobilization is useful for preparing microarrays, where various types of biological components are covalently immobilized on a plate. Because the immobilization is based on a photo-induced radical cross-linking reaction, it does not require specific functional groups on the immobilized components. Here, an aqueous solution of a photoreactive poly(ethylene glycol)-based polymer was spin-coated on a plate, and an aqueous solution of each allergen was microspotted on the coated plate and allowed to dry in air. Finally, the plate was irradiated with an ultraviolet lamp for covalent immobilization. An automated machine using these plates was developed for the assay of antigen-specific IgE. Initially, the patient serum was added to the microarray plate, and after reaction of the microspotted allergen with IgE, the adsorbed IgE was detected by a peroxidase-conjugated anti-IgE-antibody. The chemical luminescence intensity of the substrate decomposed by the peroxidase was automatically detected using a sensitive charge-coupled device camera. All the allergens were immobilized stably using this method, which was used to screen for allergen-specific IgE. The results were comparable with those using conventional specific IgE. Using this system, six different allergen-specific IgE were assayed using 10 μL of serum within a period of 20 min.Copyright © 2012 Elsevier B.V. All rights reserved.