EFFECT OF BAKERY-TYPE FOOD MATRICES AND THERMAL PROCESSING ON ELISA QUANTIFICATION OF CASHEW ALLERGEN ANA O 3

Abstract

The aim of this study was to evaluate the combined effects of cashew kernel processing, baking conditions, and food matrix type on the detectability of cashew allergens and, consequently, on the reliability of commercial ELISA kits used for allergen quantification in processed foods. The experimental design compared two baking regimes applied to muffin matrices (170 °C for 23 min and 200 °C for 18 min) with a non-thermal matrix represented by model raw bars. Cashew kernels subjected to three processing treatments (native, roasted, and blanched) were incorporated into the matrices using the geometric dilution method at nominal concentrations of 5, 10, 20, and 50 mg/kg. The results demonstrated that moisture loss during baking substantially increased the concentration of matrix solids (5.83% evaporation at 170 °C and 8.58% at 200 °C), highlighting the need for mathematical correction of allergen concentrations. Samples baked at 170 °C exhibited matrix-induced enhancement of the analytical signal and, in several cases, saturation of the assay response (ULOQ), likely due to partial protein denaturation and increased exposure of Ana o 3 epitopes. In contrast, baking at 200 °C resulted in pronounced signal suppression, with recoveries decreasing to 14.5% at 50 mg/kg of roasted cashew, a pattern consistent with extensive protein aggregation and epitope masking potentially associated with Maillard reaction products, as reported in previous studies. In the non-thermal matrix, preservation of the native protein structure led predominantly to signal enhancement and rapid saturation of the immunoassay response. Overall, the findings demonstrate that both thermal processing conditions and matrix composition substantially influence ELISA performance, potentially leading to either overestimation or underestimation of cashew allergen levels in processed food.