Species Differentiation and Quantification of Processed
Animal Proteins and Blood Products in Fish Feed Using an 8‑Plex
Mass Spectrometry-Based Immunoassay
posted on 2018-09-17, 00:00authored byAndreas
E. Steinhilber, Felix F. Schmidt, Wael Naboulsi, Hannes Planatscher, Alicia Niedzwiecka, Jutta Zagon, Albert Braeuning, Alfonso Lampen, Thomas O. Joos, Oliver Poetz
With the reintroduction of nonruminant
processed animal proteins
(PAPs) for use in aquaculture in 2013, there is a suitable alternative
to replace expensive fish meal in fish feed. Nevertheless, since the
bovine spongiform encephalopathy (BSE) crisis, the use of PAPs in
feed is strictly regulated. To date, light microscopy and polymerase
chain reaction are the official methods for proving the absence of
illegal PAPs in feed. Due to their limitations, alternative methods
for the quantitative species differentiation are needed. To address
this issue, we developed and validated an 8-plex mass spectrometry-based
immunoassay. The workflow comprises a tryptic digestion of PAPs and
blood products in suspension, a cross-species immunoaffinity enrichment
of 8 species-specific alpha-2-macroglobulin peptides using a group-specific
antibody, and a subsequent analysis by ultrahigh-performance liquid
chromatography coupled to tandem mass spectrometry for species identification
and quantification. This workflow can be used to quantitatively determine
the species origin in future feed authentication studies.