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Halbmayr-Jech et al.: Journal of AOAC International Vol. 98, No. 1, 2015 111
and strongly increased the viscosity of the extract. Hence, a clear
separation of extract aliquots was more difficult with this matrix.
This may also explain the higher CV in the homogeneity tests.
Due to the complexity of the cake recipe we cannot pinpoint a
single reason for low recovery, but a combination of the factors
mentioned is most likely. Taking this complexity into account,
the method evaluated here largely complies with the guidelines
and best practices for allergen ELISA methods (10). With an
LOD of 4.3 mg gluten/kg, it fulfills the LOD requirement of
≤10 mg/kg of Codex Alimentarius (4).
Conclusions and Recommendation
This collaborative study has shown that the G12 Sandwich
ELISA is capable of quantifying gluten in foods with an LOD
of 4.3 mg gluten/kg. This method shows good precision and
accuracy in the concentration range of most interest (20 mg/kg
and above), where it has to be decided whether a sample meets
guidelines for gluten content. Some matrix effects, especially
with the incurred chocolate cake samples, may lower recovery
as compared to spiked samples. Therefore, it may be beneficial
to occasionally check recovery by using internal reference
samples with known gluten content.
According to these results, it is recommended that the method
be accepted by AOAC as Official First Action.
Acknowledgments
We thank Peter Köhler and Katharina Schiesser for preparing
the samples and Paul Wehling and Terry Nelsen for useful
discussions on statistics.
Furthermore, we thank the following collaborators for their
participation in this study:
Guenther Augustin, Dr. Schär S.r.l, Postal, Italy
Christy Brewe, Romer Labs, Inc., Union, MO
Zsuzsanna Bugyi and Sandor Tomoszi, Budapest University of
Technology and Economics, Hungary
Dean Clarke, National Measurement Institute, Port Melbourne,
Australia
Peter Cressey, Institute of Environmental Science and Research
Ltd, Christchurch, New Zealand
Andreas Firzinger, Romer Labs Diagnostic GmbH, Tulln,
Austria
Janette Gelroth, AIB International, Manhattan, KS
Martin Hemingway, ALcontrol Laboratories, Rotherham,
United Kingdom
Rupert Hochegger, Austrian Agency for Health and Food
Safety, Vienna, Austria
Jennifer Jolly, Covance Laboratories Inc., Battle Creek, MI
Prabhakar Kasturi, Pepsico Inc., Barrington, IL
Peter Koehler, Deutsche Forschungsanstalt für
Lebensmittelchemie, Freising, Germany
Christine Poirier and Terry Koerner, Health Canada, Ottawa,
Canada
Adrian Rogers, Romer Labs UK Ltd, Runcorn, United
Kingdom
Girdhari Sharma, U.S. Food and Drug Administration, Center
for Food Safety and Applied Nutrition, Laurel, MD
Robin Sherlock, Food Allergen Control Training Analysis,
Tennyson, Australia
Carolina Sousa, Universidad de Sevilla, Spain
Steve Taylor, Food Allergy Research and Resource Program,
University of Nebraska–Lincoln, Lincoln, NE
Joanna Topping, LGC Ltd, Teddington, United Kingdom
Paul Wehling, General Mills, Inc., Minneapolis, MN
Michael Marquard, Medallion Labs, Minneapolis, MN
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Figure 2. Plot of reproducibility SD (SR) versus the global mean
observed gluten concentration for the interlaboratory study.