Food allergens are becoming increasingly threatening and are disrupting the health and social structure of a significantly large population worldwide. Proteins from mustard are among the well-recognized food allergens which affect many sensitive individuals. Many processing methods are continually being explored to reduce allergen immunoreactivity and for developing hypoallergenic foods. Cooking, germination, and fermentation have been evaluated to attenuate the immunoreactivity of food allergens. The objective of this study is to evaluate the effect of seed germination, lactic acid fermentation, and/or cooking on yellow mustard seed protein immunoreactivity (IR) (protein A) using ELISA techniques. Samples from five-day germination at 35–40 °C and three-day fermentation between 25 °C and 35 °C were evaluated. The germination and fermentation processes yielded varying reductions in the IRs of mustard proteins, with a combined yield of about 90% reduction. When complemented with further stovetop cooking, protein IR reduction was extended up to 98%, while cooking alone resulted only in about a 70% reduction. FTIR results confirmed that changes in mustard protein conformation maybe due to the unfolding and/or denaturation of mustard proteins. These processing methods are beneficial as they not only help reduce the native mustard protein IR, but also increased inherent antioxidant activities in germinated and fermented mustard seeds.