Food allergen analysis: Considerations for establishing a reference measurement system to implement EU legislation

Advancing High-Throughput MS-Based Protein Quantification: A Case Study on Quantifying 10 Major Food Allergens by LC-MS/MS Using a One-Sample Multipoint External Calibration Curve

“…Additionally, Figure D displays the variability of accuracy for the six data points on both calibration curves for the 10 allergenic food preparations. The variability of OSCC results was significantly better than traditional MSCC, mainly due to OSCC being constructed using naturally existing isotopic bodies in the selected MIRM channels, without involving any manual or instrumental operation, leading to excellent calibration curve performance. ,, In food allergen quantitative analysis, the quantified target is the total protein of the allergenic food, including allergenic and nonallergenic proteins. , Therefore, the selection of allergen ingredients or synthetic peptides as reference materials (calibrants) during the preparation of calibration curves significantly affects the accuracy of quantification results . In this study, we examined three different calibration curve preparation methods, and Figure E shows the quantitative analysis results for different calibration curves.…”

“…However, the LC-MS-based method lacks a unified calibration curve preparation strategy for food allergen quantification. , Most of the reported LC-MS-based methods commonly employed external calibration, where synthetic target peptides, allergenic proteins, or allergen ingredients serve as calibrants . Additionally, the incorporation of stable isotope-labeled (SIL) target peptides as internal standards can partially correct for matrix effects and instrument measurement variations, improving the accuracy and stability of the analytical method. , Notably, the established threshold levels for priority allergenic foods proposed by the FAO/WHO refer to the total protein content of the allergenic food, including sensitizing and nonsensitizing proteins. , Understanding this distinction is crucial for the development of allergen quantification analysis. However, many published LC-MS-based methods commonly utilize synthetic target peptides and allergenic proteins as calibrants to calculate molar quantities of peptides and derive target protein concentrations. ,, This calibration strategy, although widely applied in targeted proteomics, introduces significant measurement errors when quantifying the total protein content of allergenic foods in complex matrices .…”