Enhancing gluten detection assay development through optimization of gliadin extraction conditions

Gluten consumption can lead to severe health conditions in certain individuals, and following a strict gluten-free diet is often the only effective treatment option. Therefore, it is crucial to develop a gluten detection method that is accurate, sensitive, and specific to ensure the absence of gluten. An important aspect of developing effective gluten detection tests is the implementation of an efficient gluten extraction method. In this study, we conducted an evaluation of various buffer conditions for gliadin extraction from both heat-treated and non-heat-treated food samples. These buffer conditions included ethanol, 2-mercaptoethanol, guanidine hydrochloride, detergents, chelating agents, and deep eutectic solvents. Among the tested conditions, a combination of 2mercaptoethanol and guanidine hydrochloride demonstrated significantly higher extraction efficacy compared to most other conditions. Furthermore, we explored the use of a less toxic extraction buffer, choline chloride, which exhibited a 1.4-fold higher extraction efficiency than the combination of 2-mercaptoethanol and guanidine hydrochloride (p < 0.05). Choline chloride showed great potential as a preferred buffer for commercial gliadin extraction kits, suitable for both heat-treated and non-heat-treated food samples. Overall, our findings highlight the importance of optimizing the gluten extraction process to improve the accuracy and reliability of gluten detection methods, ultimately contributing to the development of effective tools for individuals following a strict gluten-free diet.