Background and Aims: Various guidewires are used for biliary cannulation, and each one has its own characteristics affecting its effectiveness. This study aimed to measure the basic properties and evaluate the effectiveness of a newly developed 0.025-inch guidewire for selective biliary cannulation. Methods: A total of 190 patients at five referral hospitals were randomly allocated to undergo selective biliary cannulation using the newly developed guidewire (NGW group,n= 95) or a conventional guidewire (CGW group,n= 95). The primary outcome was the selective biliary cannulation rate in naïve papillae. The secondary outcome was to measure the NGW basic properties, compare them with those of the CGW, and analyze the importance of basic property differences. Results: There were no significant differences between the groups in the baseline characteristics. The primary outcome (75.8% vs. 84.2%,p= 0.102) and adverse event rate (6.3% vs. 4.2%,p= 0.374) were similar in both groups. However, compared with the CGW group, the NGW group showed a higher number of ampulla contacts (2.58 vs. 2.02,p= 0.011) and longer cannulation time (216.5 vs. 135.1 s,p= 0.016). Furthermore, the NGW group had higher maximum friction (34.6 ± 1.34 vs. 30.2 ± 4.09), lower stiffness, and better elastic resiliency. In the multivariate analysis, a curved-tip GW (OR = 0.26, 95% CI 0.11–0.62,p= 0.002) and normal papillary shape (OR = 0.39, 95% CI 0.17–0.86,p= 0.021) were contributing factors for successful selective biliary cannulation. Conclusions: The NGW group had high friction and low stiffness, characteristics affecting biliary cannulation. Clinically, the NGW group had similar success and adverse event rates as the CGW, but they showed a higher number of ampulla contacts and longer cannulation time.