Lao People’s Democratic Republic (Lao PDR) submitted a request to Japan for 200,000 doses of expired foot-and-mouth disease (FMD) O type vaccines that were in storage for emergency use. Approximately 100,000 animals, consisting of both cattle and Asian water buffalo (Bubalus bubalus bubalis), received the same vaccine twice within one month in Xieng Khouang province in the northeast area of Lao PDR. Concurrently, the efficacy of three-month expired FMD O type vaccine (6PD₅₀O Manisa) was assessed in serum samples of 90 cattle and 31 buffalo from the field using a Liquid Phase Blocking-ELISA (LPBE) assay. Of these samples, 75 cattle (83.3%) and 24 buffalo (77.4%) were seropositive against the FMD virus (FMDV) O type before vaccination. Testing for non-structural protein (NSP) using the PrioCHECK FMD NS kit showed that many of the animals with high titers in the screening test before vaccination were FMDV-infected animals. Fifteen cattle and seven buffalo with titers 1:32 or under before vaccination exhibited high titers of antibody (1:45-1:1448) one month after the first vaccination and further increased titers (1:362-1:5792) one month after the second vaccination. Nearly all of the cattle (97.6%) had high titers to control FMD 14 months after the second vaccination. To date, no outbreak of FMD has been reported at the study site. Three-month expired FMD O type vaccines induced appropriate immune responses against FMD in both cattle and buffalo.