OMB First to Final Action Recommendation Checklist 11-12-18

ERP - GLUTEN ASSAYS: AOAC 2015.05 FAR

AOAC RESEARCH INSTITUTE AOAC OFFICIAL METHODS OF ANALYSIS (OMA) OMAMAN-19: R-Biopharm, Competitive Enzyme Immunoassay Based On The R5

Monoclonal Antibody To Determine Partially Hydrolysed Gluten In Foods Containing Wheat, Rye, And Barley Study Director: Dr. Markus Lacorn, R-Biopharm AG, An der neuen Bergstraße 17, 64297 Darmstadt, Germany/ Co- Study Director: Patricia Meinhardt, R-Biopharm Inc., 870 Vossbrink Drive, Washington, MO 63090

Are there any concerns regarding the safety of the method? ER 1 none ER 2 none ER 3 No ER 4 No concerns all is well described ER 5 No. ER 6 No Are there any concerns regarding the data manipulation, data tables, or statistical analysis? ER 1

With a %CV of 10%, I would expect only two significant figures. Also, why were only duplicate analyses performed? We routinely use triplicates and report a mean. Here, both individual values are given resulting in tables giving the impression of massive amounts of data, but such is actually not the case.

ER 2 ER 3 ER 4

units need changing

No

No concerns, perhaps that the authors can look at AOAC guidelines only - it is "nice to know" that you come to a lower LOD in the SLV/in house study with IUPAC guidelines; but would you have reported this when the LOD's with an alternative calculation to AOAC's came out with 30-40% higher LOD's? Yes. The manipulations of data to calculate spike recovery may have some questions. The estimations of the true value of the secalins in the fermented sourdough were not confirmed by an alternate established method in this as well as the spiked samples. This may result in question to the values used for calculation of spike recovery. The manuscript throughout except few places in text provides result as mg prolamin/kg. The title of the method states partially hydrolyzed (analysis) in fermented cereal- based products. The kit insert declares it as gliadin analysis. It will be useful to provide results to fit to the main objective (analysis of gluten/gliadin) of the method. The LOD of the method requires clarification mss states 5 mg/kg as LOD determined by manufacturer and they also provide alternate calculation of the LOD = 6.5 mg/Kg. Which value is the represents the method's LOD?

ER 5

ER 6 No General Comments (2) ER 1

The manuscript presented does not demonstrate the method as valid-for purpose.

ER 2 ER 3 ER 4 ER 5

N/A N/A N/A

The manuscript throughout except few places in text provides result as mg prolamin/kg. The title of the method states partially hydrolyzed (analysis) in fermented cereal- based products. The kit insert declares it as gliadin analysis. It will be useful to provide results to fit to the main objective (analysis of gluten/gliadin) of the method.

ER 6

N/A

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