The present investigation was initially undertaken to see if there exists Na⁺-K⁺ activated ATPase in the microsome fraction of the kidney. Having confirmed the presence of such an enzyme, further attempts have been made to characterize its nature and the following conclusions were obtained: (1) The ATPase activity was greatest at the Na⁺ concentration of 100 mM as well as at K⁺ concentration of 10 mM. Moreover, the ATPase activity was found to be depressed by Ca⁺⁺ in the presence of Mg⁺⁺. (2) While the ATPase activity was depressed by Ouabain, the magnitude of inhibition was greater in the Na medium than in the K medium. (3) NaCN augmented the ATPase activity whereas NaF and IAA depressed it. On the other hand, DNP had little influence on the ATPase activity. (4) Diamox, vasopressin and aldosterone had no effect while HgCl₂ markedly depressed the ATPase activity These findings indicate that the nature of ATPase isolated from the microsome fraction of the rabbit kidney is quite similar to that from other organs such as the heart and the muscle, although there are certain features specific to the type of organs.