In vitro 3-dimensional (3D) cell cultures produce valuable models that mimic 3D tissue organization and function and enhance the understanding of cell/tissue function under normal and pathological situations. Tissue function depends on the interactions between cells and the extracellular matrix; thus, effective 3D cell cultures rely on the use of appropriate extracellular matrix cues. Noticeable progress in 3D cell culture was obtained from studies with epithelial cells from organs of the female reproductive system including the mammary glands, the uterus, and the ovaries. These models show that replicating normal tissue organization in the resting phase is a prerequisite for appropriate physiological and pathological investigations. The authors' goals are to explain the importance of mimicking detailed aspects of normal epithelial organization and function, such as basoapical polarity, in 3D cell culture and to discuss how effective 3D cell culture models can lead to meaningful applications in reproductive biology.