A major concern in prion disease transmission is the spread of the disease agent by means of secretions and excretions. We analyzed buccal swab samples obtained from preclinical scrapie-infected sheep by concentrating the collected prions on silicon dioxide, followed by amplification by serial protein misfolding cyclic amplification. Data clearly demonstrate that prions are present in buccal swab samples from sheep with a VRQ/VRQ PRNP genotype during preclinical scrapie infection. These data describe for the first time to our knowledge the secretion of prions into the oral cavity of sheep, a finding with implications for the transmission of ovine scrapie and very likely other prion diseases.