Although the use of the insulin tolerance test (ITT) for the diagnosis of adult GH deficiency is well established, diagnostic peak GH cut-points for other commonly used GH stimulation tests are less clearly established. Despite that fact, the majority of patients in the United States who are evaluated for GH deficiency do not undergo insulin tolerance testing. The aim of this study was to evaluate the relative utility of six different methods of testing for adult GH deficiency currently used in practice in the United States and to develop diagnostic cut-points for each of these tests. Thirty-nine patients (26 male, 13 female) with adult-onset hypothalamic-pituitary disease and multiple pituitary hormone deficiencies were studied in comparison with age-, sex-, estrogen status-, and body mass index-matched control subjects (n = 34; 20 male, 14 female). A third group of patients (n = 21) with adult-onset hypothalamic-pituitary disease and no more than one additional pituitary hormone deficiency was also studied. The primary end-point was peak serum GH response to five GH stimulation tests administered in random order at five separate visits: ITT, arginine (ARG), levodopa (L-DOPA), ARG plus L-DOPA, and ARG plus GHRH. Serum IGF-I concentrations were also measured on two occasions. For purposes of analysis, patients with multiple pituitary hormone deficiencies were assumed to be GH deficient. Three diagnostic cut-points were calculated for each test to provide optimal separation of multiple pituitary hormone deficient and control subjects according to three criteria: 1) to minimize misclassification of control subjects and deficient patients (balance between high sensitivity and high specificity); 2) to provide 95% sensitivity for GH deficiency; and 3) to provide 95% specificity for GH deficiency. The greatest diagnostic accuracy occurred with the ITT and the ARG plus GHRH test, although patients preferred the latter (P = 0.001). Using peak serum GH cut-points of 5.1 microg/liter for the ITT and 4.1 microg/liter for the ARG plus GHRH test, high sensitivity (96 and 95%, respectively) and specificity (92 and 91%, respectively) for GH deficiency were achieved. To obtain 95% specificity, the peak serum GH cut-points were lower at 3.3 microg/liter and 1.5 microg/liter for the ITT and ARG plus GHRH test, respectively. There was substantial overlap between patients and control subjects for the ARG plus L-DOPA, ARG, and L-DOPA tests, but test-specific cut-points could be defined for all three tests to provide 95% sensitivity for GH deficiency (peak GH cut-points: 1.5, 1.4 and 0.64 microg/liter, respectively). However, 95% specificity could be achieved with the ARG plus L-DOPA and ARG tests only with very low peak GH cut-points (0.25 and 0.21 microg/liter, respectively) and not at all with the L-DOPA test. Although serum IGF-I levels provided less diagnostic discrimination than all five GH stimulation tests, a value below 77.2 microg/liter was 95% specific for GH deficiency. In conclusion, the diagnosis of adult GH deficiency can be made without performing an ITT, provided that test-specific cut-points are used. The ARG plus GHRH test represents an excellent alternative to the ITT for the diagnosis of GH deficiency in adults.