Based on the fact that the follicular phase of the menstrual cycle is variable in length, it has been assumed that the duration of estrogen (E2) priming could also be variable. A controversy has emerged however between studies indicating that an extended duration of E2 only treatment alters ART outcomes and others that challenged such effect. Therefore, we conducted our own analysis to further determine the impact of the duration of estrogen priming on ART outcomes following single frozen blastocyst transfers in E2 and progesterone cycles. Retrospective cohort study. Patients <40 years of age undergoing single Frozen Embryo Transfers (FET) after their first autologous IVF cycle in our center between 01/2017 and 12/2019 were analyzed. We considered only patients with a morphologically normal uterus on hysterosonogaphy and/or hysteroscopy. We excluded patients with endometrial thickness <7 mm after estrogen administration. Endometrial preparation was achieved using a variable duration of oral E2 (4mg BID) in addition to a combination of subcutaneous and vaginal progesterone. Single blastocyst transfer was performed on the 6th day of progesterone exposure. The primary endpoint was livebirth rate (LBR) and secondary endpoints included implantation (IR), clinical pregnancy (CPR) and early pregnancy loss rates as well as infant birthweight. The impact of the duration of E2 administration on LBR, CPR, IR and pregnancy loss was assessed using multivariable logistic regression models, adjusting for age, body mass index, endometrial thickness, blastocyst development day and embryo morphology grade. Effect on infant birthweight was assessed using a multivariable linear regression. The duration of E2 administration (mean: 16.3; range: 10-39days) before FET did not affect implantation (odds ratio [OR] = 0.994, 95% confidence interval CI [0.960; 1.030]), clinical pregnancy (OR= 0.99, 95%CI [0.93; 1.01]), early pregnancy loss (OR=1.05, 95% CI [0.854; 1.10]), and live birth rates (OR= 1.016, 95% CI [0.980; 1.054]). Also, the duration of estrogen exposure did not affect infant birthweight (in grams) (r= -0.04, p=0.4415). Our findings suggest that variation in the duration of E2 supplementation before progesterone initiation does not affect the outcome of single frozen blastocyst transfers, including infant birth weight.