Progesterone is pivotal to maintain pregnancy in the first trimester and low concentration (<4 ng/mL) has been associated with early pregnancy loss. Measurement of progesterone is widely used in practice to determine whether a mare needs progestin supplementation. Therefore, the objective of the present study was to determine progesterone concentration and the luteal tissue area in mares non-bred, and those bred becoming pregnant, and those failing to become pregnant. We hypothesized that pregnant mares have greater progesterone concentration than non-pregnant mares. Fifty-two cycles of mares (n = 14) were monitored by ultrasonography every other day until detection of a pre-ovulatory follicle. Then deslorelin acetate was administered to induce ovulation. Twenty-four hours later, mares were bred (∼2 billion progressively motile sperm extended in 50 mL; n = 37 cycles) or a sham-bred (50 mL of extender; n = 15 cycles). Ovulation was confirmed and number of corpora lutea and the luteal tissue area were recorded daily until 10-days post-ovulation. Progesterone concentration was assessed daily from the day of the ovulation up to 10-days post-ovulation. Pregnancy diagnosis was carried out at 10- and 13-days post-ovulation. Of the bred mares, 20 of them became pregnant and 17 did not. Data were analyzed with a mixed model, Tukey's test as post-hoc, and Pearson's coefficient of correlation. Progesterone concentration and luteal tissue area varied with time (P = .001) but not with group (P > .05). Multiple ovulations were associated with greater progesterone concentration and luteal tissue area (P = .0001). There was a moderate positive association between the number of ovulations and luteal tissue area (r = 0.54; P = .0001). The lack of change in the progesterone concentration and luteal tissue area between bred and non-bred mares suggests that horse seminal plasma does not affect luteal function in mares. As all mares had progesterone above 4 ng/mL after 5-days post-ovulation; it is possible that if mares with abnormal progesterone concentration were used, the results could have been different. In conclusion, pregnancy was not associated with greater progesterone concentration or luteal tissue area.