Allergen Microarrays for In Vitro Diagnostics of Allergies: Comparison with ImmunoCAP and AdvanSure

Abstract

Background: In vitro detection of the allergen-specific IgE antibody (sIgE) is a useful tool for the diagnosis and treatment of allergies. Although multiple simultaneous allergen tests offer simple and low-cost screening methods, these platforms also have limitations with respect to multiplexibility and analytical performance. As an alternative assay platform, we developed and validated a microarray using allergen extracts that we termed "GOLD" chip.

Methods: Serum samples of 150 allergic rhinitis patients were used in the study, and the diagnostic performance of the microarray was compared with that of AdvanSure (LG Life Sciences, Daejun, Korea) and ImmunoCAP (Phadia, Uppsala, Sweden). Standard IgE samples were used for the quantitative measurement of sIgEs.

Results: The microarray-based assay showed excellent performance in the quantitative measurement of sIgEs, demonstrating a linear correlation within the range of sIgE concentrations tested. The limit of detection (LOD) was lower than 0.35 IU/mL, which is the current standard for the LOD cut-off. The assay also provided highly reproducible sets of data. The total agreement percentage of positive and negative calls was 92.2% compared with ImmunoCAP. Moreover, an outstanding correlation was observed between the microarray and the ImmunoCAP results, with Cohen's kappa and Pearson correlation coefficient values of 0.80 and 0.79, respectively.

Conclusions: The microarray-based in vitro diagnostic platform offers a sensitive, reproducible, and highly quantitative method to detect sIgEs. The results showed strong correlations with that of ImmunoCAP. These results suggest that the new allergen microarray can serve as a useful alternative to current screening platforms, ultimately becoming a first-line screening method.

Keywords: Allergen microarray; Allergic rhinitis; Allergy; In vitro diagnostics; Performance evaluation; sIgE.

Fig. 1. Schematic representations of the GOLD chip and the detection of allergen-specific IgEs. (A) Allergens are spotted onto hydrophilic polymer-coated glass slides in triplicate for immobilization. The assay is performed in a silicon chamber and the results are analyzed by fluorescence scanning. (B) Schematic representation of the array. The circles represent the spotting positions of the allergen extracts (in triplicate). (C) The allergens are immobilized via covalent bond formation between the NHS ester on the activated slide and an amine on the protein surface.

Fig. 2. Analysis of allergen microarray-based assays. (A) The concentration of allergen-specific IgE (sIgE) was measured quantitatively using standard human IgEs. (B) Quantitative evaluation of varying amounts of sIgE for D. pteronyssinus in human sera. (C) Quantitative evaluation of varying amounts of sIgE for D. farinae in human sera. (D) Sera from four patients displaying different sIgE profiles.

Fig. 3. Correlation analysis of three in vitro diagnostic platforms: GOLD chip, ImmunoCAP, and AdvanSure. The sera of 150 patients with allergy were screened for seven of the most common allergens, and the amount of allergen-specific IgE (sIgE) measured by each platform was plotted for Pearson's correlation analysis.