ImmunoCAP ISAC E112i (ISAC) and Allergy Explorer 2 (ALEX²) detect specific immunoglobulin E (IgE) reactivity. Both multiplex assays contain molecular allergens and ALEX² additionally includes allergen extracts and inhibitors that block the binding of IgE to cross-reacting carbohydrate determinants (CCDs). This study aimed to compare the performance of ISAC and ALEX² by determining the IgE reactivity against allergen extracts and/or allergen components and by using qualitative, semiquantitative, and quantitative analyses of all comparable allergen components in sera from 216 participants recruited in South Tyrol/Italy. For extract sensitization in ALEX², the analysis revealed negative corresponding allergen components in 18.4% and at least one positive corresponding allergen component in 81.6% of all cases. For ISAC, the corresponding results were 23.5% and 76.5% of cases, respectively. The ALEX² CCD inhibitor eliminated CCD-positive signals detected by ISAC in 88.5% of cases. Based on sensitization values of 0.3–14.9 ISU or kU_A/L, there was good agreement between ALEX² and ISAC, although ALEX² showed higher values than ISAC. The addition of allergen-extract tests in ALEX² resulted in the detection of more sensitizations than with corresponding allergen components alone. In the range of <15 ISU or kU_A/L, ALEX² may be more effective in detecting sensitizations.

The ALEX²-test (MacroArray Diagnostics, Vienna, Austria) is a diagnostic multiplex IgE-test for the simultaneous detection of IgE to 178 allergens and 117 extracts, in addition to total IgE. Test results from more than 90 countries are stored on a GDPR-compliant cloud server for backup, customer support, and continuous postmarket surveillance. To improve the coverage of exposomes on a global scale and to further increase the sensitivity of the test, the allergen panel was updated from ALEX² to ALEX³. By mid-2023, when ALEX³ was designed, almost 400,000 real-world ALEX² test results were available. Analysing prevalences and average sIgE-levels of individual allergen preparations, coverage of extracts by components, and co-reactivity of members of the same allergen family provided a rationale for updating the array. In parallel, based on the scientific literature and clinical studies, new allergens were selected. On ALEX³, 218 allergens and 82 extracts will be represented, including 52 new allergens. Allergen preparations with low prevalence and clinical relevance, as well as redundant allergens and extracts, were discontinued. New allergens encompass, e.g., cyclophilins, alpha-gal, and additional markers from respiratory and food allergen sources. Using a large dataset of ALEX² test results exemplifies the targeted, data-driven improvement of a diagnostic IgE-macroarray.

Oral food challenge (OFC) remains the gold standard for the diagnosis of food allergies. However, this test is not without risks, given that severe allergic reactions can be triggered while it is conducted. The purpose of this study is to identify potential demographic variables, clinical characteristics of the patients and biomarkers that may be associated with severe reactions during the hazelnut oral challenge test. The sample included 22 children allergic to hazelnut who underwent a tree nut skin prick test (SPT), specific IgE (sIgE) to hazelnut, component-resolved diagnosis (CRD) with different hazelnut allergens (Cor a 1, Cor a 8, Cor a 9, Cor a 11, Cor a 14), and a single-blind placebo-controlled challenge with hazelnut. A statistically significant relationship was found between the severity of the reaction and the highest values of sIgE to hazelnut, Cor a 11 and Cor a 14, cumulative symptom-triggering dose and sunflower seed sensitization. The use of the CRD is a useful tool to identify patients at higher risk of developing a severe reaction. In this pediatric population sample from Spain, storage proteins were confirmed to be most involved in hazelnut allergy and the development of severe reactions.