The xMAP Food Allergen Detection Assay (xMAP FADA) can simultaneously detect 15 analytes (14 food allergens, plus gluten) in one analysis. The xMAP FADA typically employs two antibody bead sets per analyte, providing built-in confirmation that is not available with other antibody-based assays. Before an analytical method can be used, it is important to assess its reliability when conditions of the assay procedure are altered. This study reports the effects on assay performance associated with incubation temperature and varying amounts of bead cocktail, and also detection antibody and β-mercaptoethanol concentrations in the reduced-denatured extraction buffer. The analysis of buffered-detergent extracts displayed lower responses at 22°C compared to 37°C, while temperature had no effect on the analysis of reduced-denatured extracts. Changes in β-mercaptoethanol and detection antibody concentrations only displayed an effect on the detection of milk in the reduced-denatured extracts. A slight change in the measured bead count was observed when one-fourth the bead cocktail was used, and displayed a large decrease when one-eighth of the recommended amount was used, but this number (≥25) was still sufficient to provide reliable results. Overall, the xMAP FADA displayed an excellent robustness towards changes in the assay procedure, as may inadvertently occur.