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Background Pregnancy represents a unique immunological adaptation, balancing maternal immune responses to protect the mother and the semi-allogeneic fetus. Since this biological process remains underexplored, this longitudinal study aims to characterise immune cell populations and phenotypes (natural killer (NK) cells, monocytes, B and T lymphocytes, regulatory T (Treg) cells, helper T (Th) cells) throughout the three trimesters of healthy pregnancies.Methods A cohort of 30 women attending routine low-risk prenatal consultations was enrolled in this study. Peripheral blood samples were used to perform haemogram analysis and multiparametric flow cytometry to assess immune cell frequencies at three stages: first (T1), second (T2) and third (T3) trimesters.Results Significant changes were observed in haematological parameters, including a decrease in red blood cells and haemoglobin from T1 to T3 (p<0.001) and an increase in monocytes in both percentage and absolute count (p<0.05). Flow cytometry revealed a rise in T lymphocytes, particularly CD8 + T cells, from T1 to T3 (p<0.001) and a significant increase in Th2 cells (p<0.001). Conversely, the relative frequency of Th17 and Th17.1 cells significantly decreased (p<0.05). Treg cells also exhibited notable changes, with a decrease in memory Tregs and activated Tregs over the course of pregnancy (p<0.05). Additionally, NK cells showed a shift, with an increase in CD56bright NK cells (p<0.001) and a decrease in CD56dim NK cells (p<0.001).Conclusions These findings enhance our understanding of immune regulation during pregnancy and can inform future strategies for maternal-fetal medicine and high-risk pregnancy management.