Introduction: Among chronic diseases, knee osteoarthritis (KOA) is a joint disease that causes important progressive alterations in the articular and periarticular structures, including synovial inflammation. Exercise has been suggested as an intervention to KOA individuals, and studies suggest that whole-body vibration (WBV) exercise decreases pain levels and favours the functionality of KOA individuals. Objective: The aim of the present study is to analyze the acute effects of WBV exercise on pain levels, functionality (Timed Up and Go (TUG test), anterior trunk flexion (ATF)), and rating of exertion of elderly obese KOA individuals. Methods: Thirty-seven individuals with KOA were allocated to a WBV exercise group (WBVEG), n = 19 (15 females/4 males), and a control group (CG), n = 18 (15 females/3 males). WBVEG performed one session of WBV exercise (11 min, using 5 Hz, 2.5, 5.0, and 7.5 mm, 0.12, 0.25, and 0.37 g). Three bouts were performed (working time of 3 min and rest time of 1 min) using a side-alternating vibrating platform (VP). The same position was used in CG; however, the VP was turned off and there was equipment coupled to the VP that emitted a sound like the vibrations. The pain level was evaluated using a visual analog scale (VAS). Functionality was evaluated with a TUG test and ATF. The rating of subjectively perceived exertion was measured with the category ratio CR-10 (BORG Scale CR-10), Results: A reduction of pain levels in WBVEG after the intervention (p = 0.001) and intergroups (p = 0.041) was found. A decrease of TUG test time in both groups (p = 0.001) and intergroups (p = 0.045) was found, while no statistical changes were observed in the Borg Scale score. Significant improvements of flexibility in both groups (p = 0.001) and intergroups (p = 0.043) were found. Conclusion: One session of WBV exercise can lead to important improvements in individuals with KOA, possibly triggered by physiological responses. However, more studies are needed, in this clinical context, to confirm these results.