BACKGROUND: Upper limb dysfunction is common after stroke, posing an important challenge for post-stroke rehabilitation. The clinical efficacy of acupuncture for the recovery of post-stroke upper limb function has been previously demonstrated. Mirror therapy (MT) has also been found to be effective. However, the effects of acupuncture and MT have not been systematically compared. This trial aims to elucidate the synergistic effects of acupuncture and MT on upper limb dysfunction after stroke. METHODS: A 2 × 2 factorial randomized controlled trial will be conducted at the rehabilitation hospitals affiliated with Fujian University of Traditional Chinese Medicine. A total of 136 eligible subjects will be randomly divided into acupuncture treatment (AT), MT, combined treatment, and control groups in a 1:1:1:1 ratio. All subjects will receive conventional treatment. The interventions will be performed 5 days per week for 4 weeks. AT, MT, and combined treatment will be performed for 30 min per day (combined treatment: AT 15 min + MT 15 min). The primary outcomes in this study will be the mean change in scores on both the FMA and WMFT from baseline to 4 weeks intervention and at 12 weeks follow-up between the two groups and within groups. The secondary outcomes are the mean change in the scores on the Visual Analogue Scale, Stroke Impact Scale, and modified Barthel index. Medical abstraction of adverse events will be assessed at each visit. DISCUSSION: The results of this trial will demonstrate the synergistic effect of acupuncture and MT on upper limb motor dysfunction after stroke. In addition, whether AT and MT, either combined or alone, are more effective than the conventional treatment in the management of post-stroke upper limb dysfunction will also be determined. TRIAL REGISTRATION: Chinese Clinical Trial Registry: ChiCTR-IOR-17011118 . Registered on April 11, 2017. Version number: 01.2016.09.1.