The plasticity of the mammary gland relies on adult mammary stem cells (MaSC) and their progenitors, which give rise to various populations of mammary epithelial cells (MEC). To face global challenges, an in-depth characterization of milk-producing animal mammary gland plasticity is required, to select more sustainable and robust dairy cows. The identification and characterization of MaSC and their progenitors will also provide innovative tools in veterinary/human medicine regarding mammary tissue damage (carcinogenesis, bacterial infections). This study aimed to determine the dynamics of mammary cell populations throughout a lactation cycle. Using mammary biopsies from primiparous lactating dairy cows at 30, 90, 150 and 250 days of lactation, we phenotyped cell populations by flow cytometry. To investigate cell lineages, we used specific cell-surface markers including CD49f, CD24, EpCAM and CD10. Two cell populations linked to milk production were identified: CD49f+/EpCAM- (y = 0.88x + 4.42, R²=0.36, p<0.05) and CD49f- / EpCAM- (y = -1.15x + 92.44, R²=0.51, p<0.05) cells. Combining immunostaining analysis, flow cytometry, daily milk production data and statistical approaches, we defined a stem cell population (CD24+ / CD49f+) and four progenitor cell populations that include bi-potent luminal progenitors (CD24- / CD49f+), lumino-alveolar progenitors (CD24- / EpCAM+), myoepithelial progenitors (CD24+ / CD10-) and lumino-ductal progenitors (CD49f- / EpCAM+). Interestingly, we found that the bi-potent luminal progenitors (CD24- / CD49f+) decreased significantly (p<0.05) during lactation. This study provides the first results of mammary cells lineage, allowing insight into mammary cell plasticity during lactation.