title: Sensitive and highly specific quantitative real-time PCR and ELISA for recording a potential transfer of novel DNA and Cry1Ab protein from feed into bovine milk.
abstract: To address food safety concerns of the public regarding the potential transfer of recombinant DNA (cry1Ab) and protein (Cry1Ab) into the milk of cows fed genetically modified maize (MON810), a highly specific and sensitive quantitative real-time PCR (qPCR) and an ELISA were developed for monitoring suspicious presence of novel DNA and Cry1Ab protein in bovine milk. The developed assays were validated according to the assay validation criteria specified in the European Commission Decision 2002/657/EC. The detection limit and detection capability of the qPCR and ELISA were 100 copies of cry1Ab microL(-1) milk and 0.4 ng mL(-1) Cry1Ab, respectively. Recovery rates of 84.9% (DNA) and 97% (protein) and low (<15%) imprecision revealed the reliable and accurate estimations. A specific qPCR amplification and use of a specific antibody in ELISA ascertained the high specificity of the assays. Using these assays for 90 milk samples collected from cows fed either transgenic (n = 8) or non-transgenic (n = 7) rations for 6 months, neither cry1Ab nor Cry1Ab protein were detected in any analyzed sample at the assay detection limits.
authors.affiliation: Physiology Weihenstephan, Technische Universität München, Weihenstephaner Berg 3, 85350, Freising, Germany. patrick.guertler@wzw.tum.de
authors.forename: Patrick
authors.initials: P
authors.lastname: Guertler
authors.forename: Vijay
authors.initials: V
authors.lastname: Paul
authors.forename: Christiane
authors.initials: C
authors.lastname: Albrecht
authors.forename: Heinrich H D
authors.initials: HH
authors.lastname: Meyer
mesh_terms: D000821:Animal Feed; D000818:Animals; D000918:Antibody Specificity; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000032:analysis*; D002417:Cattle; D004274:DNA, Recombinant / Q000032:analysis*; D004731:Endotoxins / Q000032:analysis*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D030841:Food, Genetically Modified; D006460:Hemolysin Proteins / Q000032:analysis*; D008892:Milk / Q000737:chemistry*; D030821:Plants, Genetically Modified; D011994:Recombinant Proteins; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D003313:Zea mays / Q000235:genetics*
chemical_list: D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D004274:DNA, Recombinant; D004731:Endotoxins; D006460:Hemolysin Proteins; D011994:Recombinant Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis