Testing for Gluten in Food and Beer: A Not So Simple Science

In our previous posts, we looked through the testing and regulation of gluten-free foods. We’d now like to take the time to dive into the science behind these tests. How do they work? What are they testing for? How do they ensure that the foods don’t contain gluten? Warning: there’s a lot of science in this one.

The Makeup of Gluten
Gluten is a protein, which are made from long chains of amino acids and act as building blocks for the human body. Wheat, barley, and rye contain gluten, while other grains, such as buckwheat and millet, contain proteins that are not gluten and therefore safe for celiacs. Gluten is actually a protein composite, made up of a prolamin (gliadin in wheat, hordein in barley, and secalin in rye), and glutelin.

Testing for Gluten: Not So Simple
There are two categories of tests currently in use: the enzyme-linked immunosorbent assay (ELISA) test and the lateral flow tests. An ELISA test is a general category of laboratory testing that first binds the substance being tested to an antigen. The pair are then bound to a surface, which creates a signal that can be measured.

ELISA tests are further broken down into two types of tests: adsorption and “sandwich” tests. This is important, because in a sandwich test, the substance must bind to two antigens, one on the solid surface and one reagent floating in the solution (“sandwiching” the substance between the two). There must be two sites, called epitopes, for the antigens to bind. In an adsorption test, the detector molecule bonds to both the substance and the surface, meaning the substance needs only one epitope. All of the tests react with the prolamin portion of gluten.

Current Methodologies
There are several methods currently in use to detect gluten in food and beer:

  1. Omega Gliadin ELISA: Sandwich ELISA which tests for omega-gliadin. This test greatly underestimates the barley hordein, and is therefore not useful for testing beer.
  2. Standard R5 ELISA: Sandwich ELISA which tests for the epitope QQPFP, which may be toxic to celiacs. This overestimates the the barley hordein. This has been validated by the Prolamin Working Group of the Codex Alimentarius Commission, which we talked about in a previous post.
  3. Fast R5 ELISA: Sandwich ELISA which tests for the same epitope as the Standard R5. This is not validated by the Prolamin Working Group.
  4. Quick Gliadin: A lateral flow test that uses a sandwich methodology and colored nanoparticles. This has not been validated.
  5. Competitive R5 ELISA: This adsorption test requires only one epitope and is therefore more appropriate for highly hydrolyzed products, such as beer. However, the measurement is expressed in gluten peptides (short chains of amino acids) rather than gluten molecules, and it is difficult to convert one measurement to the other.
  6. Mass Spectroscopy: This is not a cheap or quick test, but can be used to more accurately detect the levels and types of gluten in beer. We discussed this method in one of our previous posts.

All of these tests do not accurately test for gluten in beer at the end of the brewing process. During the brewing process, the gluten molecules are hydrolyzed, or broken down into smaller amino acid chains. These fragments may not contain the two epitopes necessary for the sandwich methods, which means they will not be detected, but they will still cause a reaction in celiacs.

Gluten Testing

The photo above illustrates the difference between sandwich ELISA (left) and adsorption ELISA (right). In sandwich ELISA, fragments of gliadin containing only one epitope are not measured bot the test (rightmost gliadin molecule). This is not a problem with adsorption tests.

Wow, that was a lot of science. The key takeaways are:

  • Several tests exist for testing for gluten in food, each of which offer benefits and weaknesses
  • No test effectively determines gluten content in beer

In a future post, we’ll talk about some best practices when it comes to testing gluten-free beer.