Application of Mass Spectrometry-Based Immunoassays for the Species- and Tissue-Specific Quantification of Banned Processed Animal Proteins in Feeds

Processed Animal Proteins (PAPs) are considered as a sustainable protein source to improve the nutritional profile of feed for livestock and aquaculture. However, the use of these proteins is strongly regulated since the bovine spongiform encephalopathy (BSE) crisis. The reintroduction of nonruminant PAPs for use in aquaculture in 2013 has driven the need for alternative analytical methods to determine the species origin as well as the tissue source (legal or not). The current official methods, light microscopy and polymerase chain reaction, do not fulfill these requirements. Furthermore, future methods need to be quantitative, because the pending zero-tolerance-concept is planned to be replaced by accurate thresholds. Here, we developed a 7-plex mass spectrometry-based immunoassay that is capable of quantifying 0.1% (w/w) ruminant PAP in feed in a tissue- and species-specific way. The workflow comprises a 2 h tryptic digestion of PAPs in suspension, an immunoaffinity enrichment of peptides, and LC–MS/MS-...