The study aimed to evaluate the viability of Lactobacillus casei CSL3 immobilized on oat flakes applied to yogurt. Oat flakes were used as the support for L. casei CSL3, and the viability of the stored biocatalyst was evaluated at 4 ℃ and −18 ℃, lyophilized and non‐lyophilized, for 60 days. Two formulations of yogurt were produced: free L. casei CSL3 (YCL) and L. casei CSL3 immobilized on oat flakes (YCLI). Physical‐chemical analyses, cellular viability, gastrointestinal tract (GIT) simulations and sensorial tests were performed. Immobilization combined with storage temperature (4 ℃) maintained the viability of the probiotic for 60 days. The YCLI presented bacterial viability at 4 ℃ on the 25th day. GIT simulation showed reduction in the gastric phase, reaching a greater concentration in the ileum for YCLI. YCLI yogurt presented high protein content and high sensory acceptance by consumers, indicating a potential for commercialization. Practical applications: Lactobacillus casei immobilized on oats can be stored for up to 60 days in refrigeration (4 ℃) or associate the freeze‐drying and storage process at −18 ℃ for longer periods.Application of a new isolate, potentially probiotic, in the production of yogurt.The incorporation of L. casei immobilized on oats increases the contraction of protein and unsaturated fatty acids, especially oleic acid. [ABSTRACT FROM AUTHOR]