Development and validation of a sensitive enzyme immunoassay for surveillance of Cry1Ab toxin in bovine blood plasma of cows fed Bt-maize (MON810) Original Research Article

The increasing global adoption of genetically modified (GM) plant derivatives in animal feed has provoked a strong demand for an appropriate detection method to evaluate the existence of transgenic protein in animal tissues and animal by-products derived from GM plant fed animals. A highly specific and sensitive sandwich enzyme immunoassay for the surveillance of transgenic Cry1Ab protein from Bt-maize in the blood plasma of cows fed on Bt-maize was developed and validated according to the criteria of EU-Decision 2002/657/EC. The sandwich assay is based on immuno-affinity purified polyclonal antibody raised against Cry1Ab protein in rabbits. Native and biotinylated forms of this antibody served as capture antibody and detection antibody for the ELISA, respectively. Streptavidin-horseradish peroxidase conjugate and TMB substrate provided the means for enzymatic colour development. The immunoassay allowed Cry1Ab protein determination in bovine blood plasma in an analytical range of 0.4–100 ng mL−1 with a decision limit (CCα) of 1.5 ng mL−1 and detection capability (CCβ) of 2.3 ng mL−1. Recoveries ranged from 89 to 106% (mean value of 98%) in spiked plasma. In total, 20 plasma samples from cows (n = 7) fed non-transgenic maize and 24 samples from cows (n = 8) fed transgenic maize (collected before and, after 1 and 2 months of feeding) were investigated for the presence of the Cry1Ab protein. There was no difference amongst both groups (all the samples were below 1.5 ng mL−1; CCα). No plasma sample was positive for the presence of the Cry1Ab protein at CCα and CCβ of the assay.