A wee bit more information on gluten testing…

Question: Why might sandwich ELISAs underestimate the gluten content of food products containing hydrolyzed gluten?

Answer: Sandwich ELISAs require two epitopes or antibody binding sites to work. For example, the sandwich R5 ELISA is based on the R5 monoclonal antibody to the epitope QQPFP (amino acid sequence glutamine-glutamine-proline-phenylalanine-proline). So for the sandwich R5 ELISA to work, two QQPFP epitopes are required. When gluten protein has been hydrolyzed or broken into smaller protein fragments, the resulting peptides may no longer contain two epitopes or antibody binding sites. If a sandwich ELISA is used to assess the gluten content of a product containing hydrolyzed gluten (e.g., malt extract), gluten content may be underestimated.

Consider the following protein where “QQPFP” represents the epitope and “a” represents other amino acids:


If this protein undergoes hydrolysis, the following three fragments may result:

1. aaaaaQQPFP

2. aaaaaaaaaaaaaaaQQPFPaaaQQPFP

3. aaaaaaQQPFP

The sandwich R5 ELISA would be unable to measure the first or the third protein fragments because these peptides contain only 1 QQPFP epitope. Only the second protein fragment would be measured by the sandwich R5 ELISA. When only one epitope or antibody binding site is available (which may be the case with hydrolyzed ingredients), a competitive ELISA (e.g., competitive R5 ELISA) should also be used for gluten analysis.

Hopefully this helps to clarify things! If not, let me know.

Copyright © 2010 by Tricia Thompson, MS, RD

Sandwich ELISAs and Hydrolyzed Gluten