Octapeptide repeats are 24 bp long repeat segments in the PrP gene that may be present in different species in a different number of repetitions. The aim of the present study was to determine the prevalence of this polymorphism in the PrP gene of two Slovak autochtonous cattle breeds, namely Slovak spotted cattle (n = 89) and Slovak Pinzgau (n = 195). In the study's cohort of animals, repletion numbers 6 and 5 were found for the octapeptide repeats. We found 6/6 homozygous animals to be most prevalent (96.1%); the remaining animals were 5/6 heterozygous (3.9%). We did not observe any 5/5 homozygous animals in the studied group. This analysis included only a small group of animals. However, the results suggest that animals with six octapeptide repeats are dominant in these Slovak cattle breeds.