AOACRIGlutenMethods-2017Awards

H albmayr -J ech et al .: J ournal of AOAC I nternational V ol . 98, N o . 1, 2015  103

FOOD COMPOSITION AND ADDITIVES

Gluten in Rice Flour and Baked Rice Products by G12 Sandwich ELISA: First Action 2014.03 E lisabeth H albmayr -J ech Romer Labs Division Holding GmbH, Technopark 1, 3430 Tulln, Austria A drian R ogers Romer Labs UK Ltd, Block 5, The Heath Technical and Business Park, Runcorn, Cheshire WA7 4QX, United Kingdom C lyde D on Foodphysica, Vogelwikke 12, 6665 HP Driel, The Netherlands M ichael P rinster Romer Labs Inc, 1301 Stylemaster Dr, Union, MO 63084-1156 Collaborators: G. Augustin; C. Brewe; Z. Bugyi; D. Clarke; P. Cressey; A. Firzinger; J. Gelroth; M. Hemingway; R. Hochegger; J. Jolly; P. Kasturi; P. Koehler; T. Koerner; M. Marquard; C. Poirier; A. Rogers; G. Sharma; R. Sherlock; C. Sousa; S. Taylor; S. Tomoszi, J. Topping; P. Wehling

done in 60 min. Ready-to-use standards of the ELISA test kit are calibrated against the Working Group on Prolamin Analysis and Toxicity (WGPAT) gliadin standard material and cover a range from 4 to 200 mg gluten/kg sample ( see Figure 1). The preparation of ready-to-use standards was described at Halbmayr- Jech et al. (3). Single-laboratory validation (SLV), performed by Romer Labs UK Ltd in May 2011, determined an LOD of 2 mg gluten/kg sample and an LOQ of 4 mg gluten/kg sample ( see Table 1) as well as a recovery rate ranging from 90 to 145% ( see Table 2) for the Gluten G12 Sandwich ELISA assay. Coefficient of variation for repeatability and lot-to-lot variation (reproducibility) was 15% or less determined within the SLV ( see Tables 3–5). The AgraQuant Gluten G12 kit furthermore produced results similar to those assigned values for the current Codex type I approved R5 Mendez method in three Food Analysis Performance Assessment Scheme (FAPAS) rounds in 2011 ( see Table 6). The Gluten G12 Sandwich ELISA assay has been evaluated in a collaborative study with 20 participants. The main target for an allowable immunogenic gluten method according to the Codex Alimentarius is that it should have a detection limit of 10 mg/kg or below (4). This paper reports the findings of the collaborative study and discusses the results in relation to current thresholds (20 mg/kg) for gluten-free products. Study Design The study was conducted on 12 different food samples prepared in the laboratory of the Deutsche Forschungsanstalt für Lebensmittelchemie, Freising, Germany. Blind-coded samples in duplicate, ELISA test kits including extraction solution, method instructions, and result reporting sheets were sent to all participating laboratories. Collaborative Study

Received August 5, 2014. The method was approved by the Expert Review Panel for Food Allergens-Gluten as First Action. The Expert Review Panel for Food Allergens-Gluten invites method users to provide feedback on the First Action methods. Feedback from method users will help verify that the methods are fit for purpose and are critical to gaining global recognition and acceptance of the methods. Comments can be sent directly to the corresponding author or methodfeedback@aoac.org. Corresponding author’s e-mail: elisabeth.halbmayr@romerlabs.com Copyright permission has been received by Cereal Foods World to reprint the tables. DOI:10.5740/jaoacint.14-197 antibody utilized in the test kit binds to the celiac toxic amino acid sequence QPQLPY and related sequences in rye and barley (1,2). A homogenized sample is extracted with ethanol and a proprietary extraction solution containing reducing agents. The gluten determination is based on a microtiter plate coated with specific monoclonal G12 antibody. Gluten is detected with a peroxidase-labeled G12 antibody. The determination can be The Protein and Enzymes Technical Committee of American Association of Cereal Chemists initiated a collaborative study to confirm whether the G12 antibody-based sandwich ELISA test kit is able to detect gluten in the lower mg/kg (ppm) level. Twenty laboratories investigated 24 heat-treated and non-heat-treated blind-coded samples with incurred gluten levels up to 100 mg/kg. The method has been validated for testing foods to conform to the defined Codex thresholds for gluten in gluten-free products at less than 20 mg gluten/kg. The collaborative study showed that low levels of gluten could be detected by G12 Sandwich ELISA with reproducibility RSD R of 32% and repeatability RSD r of 16%. Incurred samples showed a recovery between 62 and 135%. It is recommended that the method be accepted by AOAC as Official First Action. A graQuant ® Gluten G12 is a sandwich ELISA for quantification of gluten from wheat, rye, barley, and cross-bred varieties in various foodstuffs. The G12

Collaborators

The collaborative study was coordinated by Clyde Don, Foodphysica, Driel, The Netherlands. Twenty laboratories from the food producing industry, universities, governments, contract