Different routes of natural progesterone supplementation have been tried as luteal phase support in infertility treatments. Orally administered progesterone is rapidly metabolized in the gastrointestinal tract and its use has proved to be inferior to i.m. and vaginal routes. Progesterone i.m. achieves serum progesterone values that are within the range of luteal phase and results in sufficient secretory transformation of the endometrium and satisfactory pregnancy rates. The comparison between i.m. and vaginal progesterone has led to controversial results as regards the superiority of one or the other in inducing secretory endometrial transformation. However, there is increasing evidence in the literature to favour the use of vaginal progesterone. Vaginally administered progesterone achieves adequate endometrial secretory transformation but its pharmacokinetic properties are greatly dependent on the formulation used. After vaginal progesterone application, discrepancies have been detected between serum progesterone values and histological endometrial features. Vaginally administered progesterone results in adequate secretory endometrial transformation, despite serum progesterone values lower than those observed after i.m. administration, even if they are lower than those observed during the luteal phase of the natural cycle. This discrepancy is indicative of the first uterine pass effect and therefore of a better bioavailability of progesterone in the uterus, with minimal systematic undesirable effects.