Disorders induced by cereal proteins (e.g. wheat allergy, celiac disease) are widespread in human population. Since their only effective treatment is the avoidance of the problematic proteins, patients have to be familiar with the composition of food products. For checking special foods produced for them, proper analytical methods are necessary. At the moment, in gluten analysis there are no reference methods and reference materials which model real food matrices. During the production and experimental utilisation of our previously developed reference material candidate, numerous questions emerged. As our model product is a real food matrix, interactions can be present between gluten proteins and other macro and micro components. Fat content of the baked cookies is almost 20%, which might affect the results of ELISA measurements. The detectable gluten content is significantly increasing after the defatting procedure, as a pre-treatment of samples. Moreover, baking is a common food processing step that might modify the structure of gluten proteins leading to denaturation and aggregation. In the soluble protein fraction the amount of low molecular weight proteins increases, while that of high molecular weight proteins decreases during the baking procedure.