An xMAP Food Allergen Detection Assay (xMAP FADA) was developed to meet analytical needs when responding to complaints by individuals with multiple food allergies and to address potential ambiguities associated with cross-reactive proteins. A single-laboratory validation (SLV) was conducted to examine the reliability of the xMAP FADA to detect 15 analytes individually or as part of a mixture at more than six concentrations in four foods. The xMAP FADA reliably detected the analytes despite the incurred dark chocolate and incurred baked muffins displaying recoveries of 10-20% and <60%, respectively. The high reliability for recoveries less than 60% in part reflects the statistical strength of the design of the xMAP FADA. Only crustacean, egg, and milk incurred in dark chocolate were not reliably detected using the PBST-buffered-detergent protocol. Following the reduced-denatured protocol, no problems were encountered in the detection of milk, although egg did not display a dynamic response in dark chocolate. The ruggedness of the xMAP FADA was ascertained by the ability of novice analysts to detect food allergens in baked rice cookies. Despite one analyst losing >80% of the beads and the count for one bead set dropping to seven, the assay displayed only a decrease in precision (increased standard deviations) and a change in the ratios between complementary antibody pairs.