We tested the hypothesis that 99mTc 2-methoxy-isobutyl-isonitrile (99mTc MIBI), a new radiopharmaceutical for myocardial perfusion imaging, provides accurate noninvasive detection of coronary artery disease (CAD). Imaging in patients after exercise and at rest with 99mTc MIBI was compared with imaging after exercise and redistribution with 201Tl in 12 normal subjects and 38 patients with angiographic documentation of CAD (greater than or equal to 50% diameter stenosis). We used single-photon emission computed tomography (SPECT) and computer quantitation of regional tracer distribution. The quality of reconstructed images with 99mTc MIBI judged visually was superior to that of 201Tl in 88% of all studies performed and was comparable in the others. With the limits of normal as 2.5 SD below the mean of gender-matched normal volunteers, 201Tl SPECT identified 32 and 99mTc MIBI identified 36 patients with CAD (p = 0.2). 201Tl SPECT identified 45 of 75 (60%) and 99mTc MIBI identified 59 of 75 (79%) stenosed coronary arteries (p less than 0.05). The quantitative severity of perfusion defects was similar for the two tracers. 201Tl SPECT identified 104 reversibly ischemic myocardial segments compared with 134 with 99mTc MIBI (p less than 0.05). Thus, SPECT myocardial perfusion imaging with 99mTc MIBI and computer quantitation provides an accurate method for the noninvasive detection of significant coronary artery disease. Furthermore, image quality is generally superior to 201Tl, and reversibly ischemic myocardial segments may be better identified with 99mTc MIBI.