Pregnancy diagnosis is an important part in reproduction management of ruminants. The aim of the study was to use a new method for evaluating the bPAG and cPAG in milk and blood bPAG and compare this results with the other method for pregnancy diagnosis in the cows. The study was carried out in 220 Holstein Frisian cows. Heparinised blood samples were taken from the jugular vein and stored at -20 degrees C until PAG assay by RIA. For bPAG and cPAG, RIA test, milk samples were homogenized. Pure bPAG was used as a standard tracer described by Zoli et al. (1992). The cows were diagnosed as pregnant by means of USG (Aloka SSD 210) and by rectal palpation. bPAG and cPAG concentration in milk increased after 28 day of pregnancy and showed the rapid increase near the parturition. The same results of bPAG concentration we obtained in the blood samples. The decline of bPAG concentration was faster in the milk than in the blood. The data showed that the RIA method is precise enough to measure PAG concentrations in the maternal blood and milk of cows. The data indicate that milk samples can be used for pregnancy diagnosis in cows. The sensitivity and specificity of RIA measurement of PAG are very high