Optimized In-House Basophil Activation Test for Pediatric Egg Allergy: Agreement with a Structured Composite Clinical...

Accurate diagnosis of IgE-mediated egg allergy in young children remains challenging. While basophil activation test (BAT) offers a functional readout, methodological heterogeneity and limited real-world pediatric validation have hindered its standardized use.

To develop and evaluate an optimized in-house BAT protocol for pediatric egg allergy, and to assess its agreement with structured clinical classification and its exploratory adjunctive role in diagnostically challenging cases.

Children aged 0-6 years with suspected food allergy were prospectively enrolled in a cross-sectional agreement study without longitudinal follow-up and classified by a structured composite reference standard integrating clinical history, egg-specific IgE (sIgE), documented tolerance, and expert panel consensus as egg-allergic (EA, n=28), egg non-allergic (NEA, n=68), or diagnostically indeterminate (IND, n=6). BAT was performed using stimulation with egg extract. The stimulation index (SI) was defined as the ratio of median fluorescence intensity after stimulation to that of the unstimulated control. Marker-specific thresholds were defined as the 95th percentiles of the corresponding parameters in the NEA group. Agreement with the EA/NEA classification, utility in the IND group, and correlation with sIgE were measured and evaluated.

The conjunctive CD63 SI and CD203c SI rule was selected as the primary BAT interpretation rule. In 96 EA and NEA cases, BAT showed 88.5% overall agreement as evaluated against the structured composite clinical classification (positive percent agreement 64.3%, negative percent agreement 98.5%; Cohen's κ= 0.694, 95% CI 0.530-0.858). Among 6 IND children with suggestive histories but undetectable egg-specific IgE, BAT was positive in 3 cases; however, this finding was not confirmatory of clinical allergy without oral food challenge (OFC). BAT activation parameters correlated moderately with sIgE in the overall cohort (r=0.4238-0.5126, all p<0.001), but not within the EA group alone.

This optimized in-house BAT, using a CD63 SI and CD203c SI conjunctive rule, showed substantial agreement with the structured composite clinical classification and provided functional evidence in seronegative, clinically suspected cases. However, without OFC confirmation, a positive BAT cannot establish clinical egg allergy. BAT may serve as an adjunctive functional test when first-line assessments are discordant or inconclusive, but should not be considered a replacement for OFC.