Purpose To compare the effectiveness and safety of 25-and 23-gauge sutureless microincision vitrectomy surgery (MIVS) in the management of various vitreoretinal diseases. Methods Eighty-five consecutive patients undergoing sutureless vitrectomy during January to April 2008 were randomized to either 25-or 23-gauge MIVS. Data collected prospectively included best-corrected visual acuity (BCVA), operation time, postoperative visual recovery, postoperative anterior segment change, and complications. Results The most common indications for MIVS were macular hole, macular pucker, vitreous haemorrhage, and diabetic traction retinal detachment. Mean operation times of the 25-gauge (n ¼ 38) and 23-gauge groups (n ¼ 47) were 33.68 and 34.47 min, respectively (P ¼ 0.942). Mean BCVA improved significantly in both the 25-and 23-gauge groups when measured 3-month postoperatively. There was no between-group difference in either the degree or the rate of postoperative visual recovery. Seven patients in the 23-gauge group, compared with three in the 25-gauge group, required suturing of sclerotomy at the end of the surgery. No patients in either group developed postoperative wound leakage or endophthalmitis. Conclusion Our prospective study suggests that, within the limited indications, both 25-and 23-gauge MIVS are equally effective, with similar safety profiles. Gauge selection thus may be made according to a surgeon's preference and the availability of appropriate instruments.