Background and Purpose Lesions on diffusion-weighted imaging (DWI) occasionally appear on follow-up magnetic resonance imaging (MRI) among initially DWI-negative but clinically suspicious stroke patients. We established the prevalence of positive conversion in DWI-negative stroke and determined the clinical factors associated with it.Methods This retrospective, observational, single-center study included 5,271 patients hospitalized due to stroke/transient ischemic attack (TIA) in a single university hospital during 2010 to 2017. Patients without initial DWI lesions underwent follow-up DWI imaging as a routine practice. Adjusted hazard ratios (aHRs) for recurrent stroke risk according to positive conversion were determined using Cox proportional hazard regression. Adjusted odds ratios (aORs) and 95% confidence intervals (CIs) for positive conversion among initially DWI-negative patients were estimated.Results In total, 694 (13.2%) patients (mean±standard deviation age, 62.9±13.7 years; male, 404 [58.2%]) were initially DWI-negative. Among them, 22.5% had positive-conversion on follow-up DWI. Positive conversion was associated with a higher risk of recurrent stroke (aHR, 3.12; 95% CI, 1.56 to 6.26). Early neurologic deterioration (aOR, 15.1; 95% CI, 5.71 to 47.66), atrial fibrillation (aOR, 6.17; 95% CI, 3.23 to 12.01), smoking (aOR, 3.76; 95% CI, 2.19 to 6.63), pre-stroke dependency (aOR, 1.62; 95% CI, 1.15 to 2.27), objective hemiparesis (aOR, 4.39; 95% CI, 1.90 to 10.32), longer symptom duration (aOR, 2.17; 95% CI, 1.57 to 3.08), high cholesterol (aOR, 4.70; 95% CI, 1.78 to 12.77), National Institutes of Health Stroke Scale score (aOR, 1.44; 95% CI, 1.08 to 1.91), and high systolic blood pressure (aOR, 1.01; 95% CI, 1.00 to 1.02) were associated with a higher incidence of lesions with delayed appearance. Regarding the location of lesions on follow-up DWI, 34.6% and 21.2% were in the cortex and brainstem, respectively.Conclusions In DWI-negative stroke/TIA, positive conversion is associated with a higher risk of recurrent stroke. DWI-negative stroke with factors related to positive conversion may require follow-up MRI for a definitive diagnosis.