There were analyzed the basic chemical composition, proportion of fatty acids, and cholesterol content in milk samples obtained from four breeds of cows, i.e. Polish Holstein-Friesian – Black and White variety – PHF HO (269 samples) and Red and White variety – PHF RW (302), Simmental – SM (240), and Jersey – JE (335). It was proved that in the intensive husbandry technology, the PHF HO cows had the highest daily milk yield (25.8 kg per day on average). The SM cows (representing a combined type of use) obtained a slightly lower milk yield (23.2 kg) together with a very advantageous protein to fat ratio (0.86). Milk from the SM cows had significantly the highest (p ≤ 0.05) content of short- and medium chain saturated fatty acids (SFAsmc), while milk from PHF HO and JE cows was characterized by a significantly (p≤0.01) higher proportion of long-chain saturated fatty acids (SFAlc). The content of CLA in milk from all the cow breeds analysed was rather low with significantly the highest percent level in milk from JE cows (0.35 %, at p ≤ 0.01) and SM cows (0.33 %, at p ≤ 0.01), and the lowest in milk from the PHF HO cows (0.23 %). The highest content of cholesterol (25.39 mg/100 ml) was reported in milk from the PH FEW cows, and the lowest in milk from the Jersey cows (15.71 mg/100 ml). The stage of lactation in that (intensive) livestock farming had no significant effect on the proportion of fatty acids and cholesterol content in milk. While comparing the simultaneous effect of breed and stage of lactation, significant interactions were found (p ≤ 0.01) among the daily milk yield, percent level of fat, and dry matter (p ≤ 0.01), and among the protein and lactose contents (p ≤ 0.05).