Chemical exchange saturation transfer (CEST) is an emerging MRI contrast mechanism that is capable of noninvasively imaging dilute CEST agents and local properties such as pH and temperature, augmenting the routine MRI methods. However, the routine CEST MRI includes a long RF saturation pulse followed by fast image readout, which is associated with high specific absorption rate and limited spatial resolution. In addition, echo planar imaging (EPI)-based fast image readout is prone to image distortion, particularly severe at high field. To address these limitations, we evaluated magnetization transfer (MT) prepared gradient echo (GRE) MRI for CEST imaging. We proved the feasibility using numerical simulations and experiments in vitro and in vivo. Then we optimized the sequence by serially evaluating the effects of the number of saturation steps, MT saturation power (B1), GRE readout flip angle (FA), and repetition time (TR) upon the CEST MRI, and further demonstrated the endogenous amide proton CEST imaging in rats brains (n = 5) that underwent permanent middle cerebral artery occlusion. The CEST images can identify ischemic lesions in the first 3 hours after occlusion. In summary, our study demonstrated that the readily available MT-prepared GRE MRI, if optimized, is CEST-sensitive and remains promising for translational CEST imaging.