201T1-99mTc subtraction parathyroid scintigraphy assisted by computer (CASPS) was performed in 59 patients with suspected hyperparathyroidism (HPT). There were 32 patients operated on and 40 hyperfunctioning parathyroid glands histologically proven on 33 surgical explorations. Positive rate of scintigrams in surgically explored patients was preoperatively 66% (21/32) and retrospectively 78% (25/32). In seven of surgically explored patients, ten hyperfunctioning parathyroid glands could not be recognizable retrospectively. True positive rate of preoperative and retrospective localization as to each hyperfunctioning parathyroid glands were 53% (21/40) and 75% (30/40), respectively. There was a close but not statistically significant correlation between scintigraphic localization and weight of the hyperfunctioning parathyroid glands. However, visualization of hyperfunctioning parathyroid glands was statistically significant (p less than 0.05) in X2-test in correlation between ones of less and more than 0.500 g in weight. Plain 201T1-image delineated only one of 15 hyperfunctioning parathyroid glands which were less than 0.500 g in weight and subtraction image six. This result was also statistically significant (p less than 0.05) in X2-test. Subtraction technique was proved to be essential for delineating hyperfunctioning parathyroid glands which were especially less than 0.500 g in weight. There are two adenoma which concentrated 99mTc-pertchnetate rather than 201T1 chloride. Subtraction images of these cases was not useful for positive delineation and showed an area of a decreased radioactive distribution corresponding to the presence of adenoma located at the left anterior mediastinum. However, subtraction caused false positive localization in a few cases. Quantitatively analytic method on hot regions demonstrated by subtraction, which could be helpful for avoiding false negative and false positive estimation, has been developed. Results by 201T1-99mTc subtraction parathyroid scintigraphy is not necessarily satisfactory. However, this is noninvasive and is still a challenging method for preoperative localization of hyperfunctioning parathyroid glands rather than a ultrasonography and CT scan.