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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

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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 μL−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.

Schematic formats for quantitative real-time PCR and ELISA for the quantification of cry1Ab DNA and Cry1Ab protein

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Acknowledgments

This study was funded by the Bavarian State Ministry of Nutrition, Agriculture and Forestry. The authors thank E. Englberger and T. Janke for their assistance. The staff at the Physiology Weihenstephan and the Bavarian State Research Center is gratefully acknowledged.

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Author notes

  1. Christiane Albrecht

    Present address: Institute of Biochemistry and Molecular Medicine, University of Bern, Buehlstr. 12, 3012, Bern, Switzerland

Authors and Affiliations

  1. Physiology Weihenstephan, Technische Universität München, Weihenstephaner Berg 3, 85350, Freising, Germany

    Patrick Guertler, Vijay Paul, Christiane Albrecht & Heinrich H. D. Meyer

Authors
  1. Patrick Guertler
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  2. Vijay Paul
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  3. Christiane Albrecht
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  4. Heinrich H. D. Meyer
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Corresponding author

Correspondence to Patrick Guertler.

Additional information

Patrick Guertler and Vijay Paul have contributed equally to this study.

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Guertler, P., Paul, V., Albrecht, C. et al. 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. Anal Bioanal Chem 393, 1629–1638 (2009). https://doi.org/10.1007/s00216-009-2667-2

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  • DOI: https://doi.org/10.1007/s00216-009-2667-2

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