Background: Autogenous arteriovenous fistula (AVF) is the preferred vascular access mode. However, the earliest possible time for AVF puncture and whether premature puncture affects the AVF patency rate remain unclear. Methods: In this multicenter retrospective cohort study, adult uremic patients who underwent AVF surgery for the first time at Taizhou Hospital or Enze Hospital of Zhejiang Province between September 1, 2018 and August 31, 2021 were enrolled. All patients were followed up for 1 year after puncture, and the status of fistula establishment and puncture, subsequent patency, loss to follow-up, renal transplantation, conversion to peritoneal dialysis, abandonment of the fistula, and death, were recorded. Results: A total of 465 patients with AVFs were included in this study, including 59 (12.7%) patients with fistulas that were cannulated within 30 days. In the early puncture group, the levels of serum creatinine and urea nitrogen were higher, while the levels of hemoglobin and albumin were lower, suggesting that these patients needed urgent dialysis. Furthermore, the rate of non-cuffed catheter use was higher, while the rate of cuffed catheter use was lower, and femoral vein puncture was preferred over internal jugular vein puncture. The mean duration of catheter indwelling was shorter in the early puncture group (19 vs 70 days, p < 0.001). The estimated AVF primary and cumulative functional patency at 12 months was 81.1% versus 82.3% and 98.3% versus 98.7% in the early puncture and control groups, respectively. Kaplan–Meier analysis revealed no significant difference in AVF primary and cumulative functional patency between the two groups. Conclusions: In patients with an established fistula in urgent need of hemodialysis, to avoid new catheterization, a puncture can be performed within 30 days in those with well-developed blood vessels after adequate ultrasound and clinical evaluation without affecting the patency of the fistula.