The aim of this study was to use rapid mass spectrometry (MS)-based proteomics analyses for diagnosis of Babesia canis canis infections in dogs. The study was conducted on two groups of dogs--healthy dogs and dogs infected with B. canis canis which demonstrated symptoms of babesiosis. The matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) MS technique revealed the presence of a protein fraction of 51-52 kDa in the blood serum of all the animals infected with the protozoa, which was not found in the serum of healthy dogs. The proteins are suspected to be disease markers, whereas the MALDI-TOF technique itself has high specificity and sensitivity and can be applied in analytical laboratories in the diagnosis of canine babesiosis.