Polycystin-L (PCL), the third member of the polycystin family of proteins, functions as a Ca2+-modulated nonselective cation channel when expressed in Xenopus oocytes. Polycystin-1 and -2 are mutated in autosomal-dominant polycystic kidney disease (ADPKD), but the role of PCL in disease has not been determined. In this study, an anti-peptide polyclonal antiserum was generated against the carboxyl terminal domain of human PCL and used to determine the patterns of expression and distribution of PCL by indirect immunofluorescence in both developing and adult mice. The results show that PCL is predominantly expressed in adult mouse tissues and has a more restricted pattern of expression than either polycystin-1 or -2. In the kidney, PCL expression was first detected at E16, and levels increased into adulthood. Localization of PCL was predominantly found in the apical region of the principal cells of inner medullary collecting ducts. PCL was also found in discrete cell types of the retina, testis, liver, pancreas, heart, and spleen, but it was not detected in the lung. These data in combination with evidence of PCL channel activity are crucial for elucidating the physiologic role of this novel cation channel and may shed light on the function of inner medullary collecting ducts and polycystins. The expression pattern of PCL suggests that it is unlikely to be a candidate gene for ADPKD, but it remains a potential candidate for other as yet unmapped human cystic disorders.