Carbonic anhydrase is essential for the cellular transportation of hydrogen and bicarbonate ions and plays a key role in a wide variety of physiological processes. Rainbow trout, Oncorhynchus mykiss is an important freshwater fish in aquaculture industry and is known to be one of the most susceptible species to environmental contamination. In this study, carbonic anhydrase was detected in the kidney and intestine of rainbow trout. Carbonic anhydrase was isolated from cytosolic proteins and identified by using SDS-PAGE, isoelectric focusing, and immunohistochemical methods. A specific protein band with molecular weight of 30 kDa and pI of 7.0 was detected by Western blotting. The immunohistochemical results showed that carbonic anhydrase was located at various cells in the kidney and intestine of rainbow trout.