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General comments about the method:
ER 1
The method onto itself performs as would be expected for a Competitive ELISA. This means, that the
precision of the results and sensitivity are acceptable . However, in the documentation language is at
times used that with a few minor tweaks may correct any misleading aspects (e.g., the sensitivity of the
Competitive ELISA is not better than that of the Sandwich. Instead, the Sandwich ELISA is inappropriate
for the detect not hydrolyzed gluten making it unreliable due to variability in the number of antigenic
epitopes associated with proteolysis.
ER 2
Good that all labs that participated in the study were familiar with ELISA tests, but probably would be
valuable to have experience with competitive ELISA tests. Also, the RIDASOFT cubic spline function
calculation is much different than the typical gliadin/gluten tests and requires a higher understanding of
what is going into this calculation.
ER 3
data indicates this is a fit-for-purpose method.
ER 4
Samples are extracted by a simple sample preparation and analyzed within 40 minutes.
ER 5
This configuration of the R5 ELISA format has a definite advantage over the current sandwich format and
will be a valuable tool for measuring gluten in certain matrices. This method should provide a more
accurate assessment of the gluten in products where gluten is randomly hydrolyzed.
ER 6
The competitive ELISA based on R5 antibody may be better than the sandwich format for hydrolyzed
gluten as it requires one epitope for effective binding. The accuracy of the method needs to be evaluated
towards not only the PT digest, but other hydrolyzed form with known gluten concentration as well.
ER 7
ER 8
Pros/Strengths of the Manuscript:
ER 1
The existence of competitive ELISAs is a strength and needed format for gluten methodology.
ER 2
Naturally gluten-contaminated starch syrup was included as one of the samples.
ER 3
all required sections are included
ER 4
The R5 ELISA is commercially available. The Competitive ELISA which was selected for the determination
of partially hydrolysed gluten is more sensitive than the R5 Sandwich ELISA. The Competitive Assay
quantitates beside intact prolamins, peptide fragments of prolamins from wheat (gladins), rye (secalin),
and barley (hordein).
ER 5
I found the manuscript concise and ordered appropriately.
ER 6
Good approach towards detection of hydrolyzed gluten as currently this is a big challenge.
Uses R5 antibody which is well characterized.
The in-house validation is extensive and provides useful information on the method.
The method shows good precision.
ER 7
ER 8
ERP PROFILE SUMMARIES
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