D2.85 - Analytical Performance Evaluation of Bet v 4 and Cor a 1 on IMMULITE 2000 XPi*

Panallergens such as profilins and polcalcins are highly conserved proteins found across the plant kingdom and are key drivers of cross-reactivity among pollen species. Bet v 4, a polcalcin from birch pollen, is a calcium-binding protein that represents a minor allergen but serves as a marker for broad pollen sensitization. Sensitization to Bet v 4 is generally associated with polysensitization to multiple pollen sources and may correlate with more severe respiratory disease and reduced efficacy of allergen immunotherapy. Prevalence varies geographically, ranging from 5–11% in Northern and Central Europe to up to 27% in Southern regions. Cor a 1, a PR-10 protein and Bet v 1 homologue, is the major hazelnut allergen in birch-endemic areas, accounting for up to 90% of hazelnut sensitization. IgE cross-reactivity between Bet v 1 and Cor a 1 often results in mild or clinically irrelevant food reactions, unlike seed storage proteins driving severe allergy. Together, Bet v 4 and Cor a 1 are important molecular markers for identifying cross-reactivity patterns and guiding component-resolved diagnostics in pollen and food allergy.

Limit of Blank (LoB), Limit of Detection (LoD), and Limit of Quantitation (LoQ) were determined per CLSI EP17-A2 using replicate samples across multiple reagent lots.  The highest calculated LoB/LoD/LoQ is reported. Linearity was assessed following CLSI ILA20 guidelines using serum pools diluted across the analytical range; data were analyzed by regression. Method comparison followed CLSI EP09-A3, evaluating IMMULITE 2000 XPi results against a predicate device using 50 negative and 30 positive samples; agreement was determined by concordance analysis.

LoB/LoD/ LoQ values were 0.063/0.093/0.114 kU/L for rBet v 4 and 0.062/0.083/ 0.121 kU/L for rCor a 1, respectively. Linearity was confirmed across clinically relevant ranges (0.046–94.0 kU/L) with R² ≥0.996 and slopes near unity (1.012–1.022) for Cor a 1, and 0.056–93.1 kU/L with R² ≥0.996 and slopes near unity (1.00–1.04) for Bet v 4. Method comparison demonstrated good agreement with predicate devices for both allergens, achieving 100% positive, negative, and overall agreement.

The allergens showed expected detection limits, appropriate linearity, and strong agreement with a reference method. These results confirm the assay’s sensitivity, precision, and reliability, supporting its suitability for routine clinical use in allergy diagnostics.