To evaluate the potential of superselective cavernous sinus (CS) sampling in patients with functioning pituitary adenomas, we performed direct CS sampling in 18 patients with Cushing syndrome and other functioning adenomas using a mini-catheter. Samples from the inferior petrosal sinuses (IPS) were also obtained. All of the samples from 20 CS of 10 patients with Cushing syndrome and 13 of 16 samples from CS of eight patients with other adenomas were successfully obtained. No complication occurred. In three patients, the mini-catheter was introduced to the contralateral CS through intercavernous communication, and bilateral sampling could be performed via the unilateral jugular vein. The CS/peripheral (CS/P) ratio of ACTH (mean = 66.1) was significantly greater than the IPS/peripheral (IPS/P) ratio (mean = 14.9) in Cushing disease. In other functioning adenomas, the CS/P ratio (15.8) was not significantly higher than the IPS/P ratio (4.8). Direct CS sampling offers a much higher CS/P ratio than IPS/P ratio in Cushing disease, and may provide sufficient diagnostic accuracy without CRH stimulation or bilateral simultaneous sampling.