Research into the mechanisms of prostate cancer progression has been limited by the lack of suitable in vitro systems. A hurdle in understanding the molecular genetic changes in prostate cancer has been the difficulty in establishing premalignant lesions and primary prostate tumors as in vitro cell cultures. Primary prostate epithelial cells grow for a finite life span and then senesce. Immortalization is defined by continuous growth of otherwise senescing cells and is believed to represent an early stage in tumor progression. To examine these early stages, we and others have developed in vitro models of prostate epithelial cell immortalization. Generation of primary human prostate epithelial (HPE) cells has been achieved using the serum-free condition. Retrovirus containing human telomerase reverse transcriptase (hTERT) was successfully used for the immortalization of primary HPE cells. Putative stem cell markers CD133 and CXCR4 were further identified in hTERT-immortalized primary nonmalignant and malignant tumor-derived HPE lines. In addition, an hTERT-immortalized nonmalignant HPE cell were found to retain the properties of multipotent stem cells. These in vitro prostate cell culture models should be useful for the study of carcinogenesis and of normal and cancer stem cells. Prostate cancer is the most common male cancer in the Western World and second leading cause of male cancer death in the United States [1]. The therapy most widely used against advanced disease is androgen ablation and, initially, it almost always produces objective clinical responses. However, most patients eventually relapse with ablation-resistant prostate cancer and develop metastatic disease; currently, there is no treatment that will cure progressive hormone-refractory metastatic prostate cancer. The mechanisms of progression of prostate cancer have been extensively studied, yet are poorly understood. One of the concepts that has been evolved is that cancer arises from the neoplastic transformation of normal prostate epithelial stem cells or transit amplifying cells. Understanding normal stem cells and cancer stem cells (CSCs) may provide insight into the origin of and new therapeutics for prostate cancer. However, research in this field is limited by the lack of suitable in vitro systems.