Several protocols for rest and stress myocardial perfusion imaging with 99mTc-sestamibi have been developed, each with distinct advantages and disadvantages. The various approaches have similar sensitivities and specificities for detection of coronary artery disease (CAD), but differ mainly in their ability to identify defect reversibility. The dual-isotope approach, with a rest 201Tl study and a stress 99mTc-sestamibi study, permits optimal evaluation of both stress perfusion and defect reversibility. Gated SPECT may be added to any of the protocols and aids in identifying artifacts, defining regional wall thickening and assessing ventricular function. First-pass 99mTc-sestamibi radionuclide angiography can add exercise ventricular function data to the study. Clinical trials have shown that the various protocols for 99mTc-sestamibi provide diagnostic and prognostic information comparable to that derived from traditional 201Tl imaging, with the added advantage of higher quality images and increased certainty in interpretation.