Objective: Many studies have investigated the effect of implant number on the performance of implant-supported overdentures (ISO), but few have focused on denture movement. This in-vitro study aims to investigate the influence of implant number on the movement of mandibular ISO under biting force. Method and Materials: An edentulous mandibular cast and 12 overdentures (n = 12) were fabricated, and a locator attachment system with blue nylon inserts was chosen for this study. Four implant positions were simulated: anterior midline (1-ISO), bilateral canine regions (2-ISO), anterior midline and bilateral molar regions (3-ISO), and bilateral canine and molar regions (4-ISO). After positioning an overdenture on one of these four casts, vertical loads were applied to the incisor region (40 N), the left canine (55 N), or molar (100 N) regions. The movements at the loading point and the right distal edge were measured. Significance across groups was analyzed using one-way analysis of variance and post hoc Tukey honest significant difference test. P .05 was considered statistically significant. Results: On incisor loading, the 3-ISO showed the least movement, and no significant movement was detected across the other three groups. Upon canine loading, the largest displacement appeared in the 1-ISO group, at both the loading point and the right distal edge. Additionally, overdentures anchored by three or four implants, especially the 4-ISO, showed significantly smaller movement on molar loading. Conclusion: The implant number influences the movement of mandibular ISO, and overdentures retained with three or more implants have minimal movement. Keywords: implant number, implant-supported overdenture, locator, mandibular, movement