Leptin is an important modulator of energy balance and metabolism in mammals, but for evolutionary older vertebrates like fish, the first reports on leptin expression were only recently characterized and the functional role scarcely. In this study, we demonstrated leptin immunoreactivity in liver tissue of rainbow trout (Oncorhynchus mykiss) by immunohistochemistry using three different polyclonal antibodies against mammalian leptin. Immunoreactivity was observed in hepatocytes and also in parts of the biliary system. Using Western blot, we detected an immunoreactive band of about 16 kDa in serum and visceral adipose tissue (AT) of rainbow trout. The presence of leptin in fish AT has been doubted in other studies. Besides the immunoreactivity, leptin mRNA was detected in trout AT albeit not in all animals sampled. Our observations add further evidence to the concept of AT being a source of leptin in trouts. Moreover, the cellular localization of leptin immunoreactivity in liver opens up new vistas for understanding the functional role of leptin in teleosts.