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Gluten Test Kit Methods Usda

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Testing For Glutens

  • Since this was originally posted, a new kit with a new antibody has become available. The antibody is called G12 and the kit is GlutenTox, manufactured by Biomedal. There is also an at-home version, GlutenTox Home, available at http://www.glutentox.com.

    The antibody is sensitive to wheat, barley, rye and oat and the at-home version can detect as low as 5ppm.
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Gluten Test Kit Methods Usda

  1. 1. Commercial Gluten Test Kits / Methods Jupiter Yeung, PhD Principal Scientist Food Products Association FDA Gluten Free Meeting, Aug 19, 2005
  2. 2. Gliadin Detection Methods <ul><li>Immunochemical ELISA </li></ul><ul><li>Lateral Flow Device </li></ul><ul><li>Immunoblotting </li></ul><ul><li>PCR </li></ul><ul><li>MS </li></ul><ul><ul><ul><li>MALDI-TOF </li></ul></ul></ul><ul><ul><ul><li>LC/MS/MS </li></ul></ul></ul>
  3. 3. 22 Capture Antibody Detector Antibody Signal Sandwich ELISA
  4. 4. <ul><li>Skerritt mAb </li></ul><ul><li>–  -gliadin (  -gliadin) </li></ul><ul><li>– wheat – 100% </li></ul><ul><li> rye – 120% </li></ul><ul><li> barley – 5% </li></ul><ul><li>Mendez R5 mAb </li></ul><ul><li>– QXPFP toxic epitopes of  ,  ,  ,  -gliadins </li></ul><ul><li>– Equally to wheat, rye and barley </li></ul><ul><li>Both Ab recognize raw and cooked prolamins </li></ul>Gliadin Antibodies Specificity
  5. 5. Commercial Gliadin ELISA Kits * ppm gliadin ppm gluten = ppm gliadin x 2 Converted to ppm from  g/mL Company Product LOD* LOQ* Range* Replicates Costs Neogen Alert (Qualitative) 10 1 $237/20Sampl Veratox 5 5-50 1 $465/38Sampl R-Biopharm RidaScreen 1.5 2.5 2.5-40 2 $425/96 wells RidaScreen Fast 5 5 5-40 1 $400/48 wells Tepnel Gluten Assay Kit, AOAC or High sensitivity 5 50 50-1000 2 $425/96 wells 0.5 5 5-100 2 Hallmark HAVen Gluten, AOAC or High sensitivity 5 50 50-1000 2 $425/96 wells 0.5 5 5-100 2 Diffchamb Transia Plate Gluten 5 5 5-100 2 $500/96 wells Transia Plate Prolamins 1.5 0.8 0.8-12.5 2 $500/96 wells Morinaga Wheat Protein 0.3* 3.12 3.12-20 2 $600/96 wells
  6. 6. Commercial Gliadin ELISA Kits Company Product Validation Neogen Alert Veratox R-Biopharm RidaScreen RidaScreen Fast PWG ring trial Tepnel BioKit Gluten (AOAC OM) High Sensitivity Hallmark HAVen Gluten (AOAC OM) High Sensitivity Diffchamb Transia Plate Gluten NFA, Sweden Transia Plate Prolamins Morinaga Wheat Protein NIHS, Japan
  7. 7. 0 5 10 20 50 ppm gliadin controls Gliadin ELISA
  8. 8. Lateral Flow Device (Dipstick) <ul><li>Positive can mean less than LOD </li></ul>Company Product LOD (ppm gluten) Costs R-Biopharm Rida Quick 10 $200 / 20 strips Tepnel Gluten Rapid 100 $150 / 10 strips Gluten Home 100 $ 75 / 5 strips
  9. 9. Lateral Flow Device
  10. 10. Lateral Flow Device                                                                             
  11. 11. Lateral Flow Device
  12. 12. Issues with Gluten Test Kits <ul><li>Ab has different specificities to different prolamins, e.g. Skerritt Ab </li></ul><ul><li>Various results from different extraction media: 40-60% EtOH, cocktails </li></ul><ul><li>Different standards used from different kits </li></ul><ul><li>False positives and negatives require a confirmatory test </li></ul><ul><li>Limited availability of confirmatory test to most food lab </li></ul><ul><li>No sampling plan </li></ul>
  13. 13. Extraction Media <ul><li>Aq. Alcohol (40-60% EtOH) </li></ul><ul><li>Cocktail (reducing agent + others) </li></ul><ul><li>- interaction of Ab and Ag </li></ul>
  14. 14. Matrix Effect <ul><li>Cocktail (reducing agent + others) </li></ul><ul><li>- interference with complex food matrices, e.g. milk </li></ul><ul><li>- interaction of Ab and Ag </li></ul><ul><li>Samples with high polyphenols, tannins and syrups require fish gelatin to improve recovery </li></ul>
  15. 15. <ul><li>Hasselberg et al. Proceeding of the 18 th </li></ul><ul><li>Meeting, WPGAT, 2004 </li></ul>Effects of Extraction Media
  16. 16. Effect of Partial Hydrolysis Ferre et al. Proceeding of the 18 th Meeting, WPGAT, 2004
  17. 17. Reference Materials <ul><li>IRMM-480 Wheat Gliadin </li></ul><ul><li>NIST </li></ul><ul><li>SRM 1567a Wheat Flour </li></ul><ul><li>RM 8418 Wheat Gluten </li></ul><ul><li>RM 8436, 8437, 8438, 8441a </li></ul><ul><li>Others </li></ul>
  18. 18. Confirmation Methods <ul><li>Ab-Western blot </li></ul><ul><li>PCR – qualitative, quantitative </li></ul><ul><li>MS – MALDI-TOF, LC/MS/MS </li></ul>
  19. 19. Garcia et al., Proceeding of the 18 th Meeting, WPGAT, 2004
  20. 20. Detection & Speciation Wheat Gliadins Rye Scalins Barley Hordeins Oat Avenins Ab-based + - PCR + + + + MS + + + +

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