Analytical Sciences

Abstract − Analytical Sciences, 23(5), 545 (2007).

Optimization of Critical Factors Affecting the Performance of an Allergen Chip for the Analysis of an Allergen-specific Human IgE in Serum
Min-Kyu HAN,* Moon-Hyeong SEO,* Dohoon LEE,* Seung Hyun KIM,** Hae-Sim PARK,** and Hak-Sung KIM*
*Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon 305-701, Korea
**Department of Allergy and Clinical Immunology, Ajou University School of Medicine, Suwon 442-749, Korea
A sensitive and multiplexed assay of allergen-specific human immunoglobulin E (IgE) is of great significance in the precise diagnosis of allergies. We report on the optimization of critical factors for chip-based analysis of IgE in human serum with a high reliability. Extracts of two mite species were used as model allergens, and were spotted onto a glass slide for the construction of an allergen chip. Respective allergen-specific IgE in human serum was analyzed by using biotinylated anti-human IgE and a streptavidin-Cy3 conjugate. Factors affecting the performance of the allergen chip were investigated and optimized. Especially, the effect of additives, the concentrations of biotinylated anti-human IgE and the streptavidin-Cy3 conjugate, the serum dilution factor, and the concentration of allergen extract as a capturing agent were examined in detail. Under the optimized conditions, a chip-based analysis for sera from 43 patients revealed a reliable and reproducible diagnosis of respective allergies, showing a good correlation with a conventional CAP assay.