Objective To explore the implementation of on-the-job training pathway for stroke specialist nurses under the medical alliance model and to evaluate the effectiveness of training. Methods The subjects of the study were 32 professional nurses in primary stroke center of the first People's Hospital of Yichang. We used the combination of network teaching and clinical practice to train management group of specialist nurses. Before and after the training, evaluated the professional capacity level of specialist nurses by theory examination and practicing examination. Results The qualification rate of stroke nurses' professional knowledge was increased from 37.5% (12/32) up to 100.0% (32/32), (χ²＝29.09, P<0.01), the qualified rate of the door-to-needle times raised to 86.2%(χ²＝4.23, P<0.05). Conclusions Implementing the on-the-job training path extends the professional knowledge of stroke nurses. Improving the qualification rate of DNT of the patient in medical alliance units, so more stroke victims benefit from early vascular re-canalization .