Introduction: Staphylococcus aureus is the leading biofilmforming microorganisms in orthopaedic implant infections. The biofilms formed are difficult to eradicate and resistance to antibiotics. This current study aims to determine the effectiveness of povidone-iodine; an antiseptic solution in eradicating S. aureus biofilm on stainless steel alloy. In addition to the usual Colony-Forming Unit (CFU) used for verification, Scanning Electron Microscope (SEM) is used to validate the formation and eradication of the biofilms. Materials and methods: This is an in vitro study where the biofilm is formed by inoculating clinically isolated S. aureus, incubated for 24 hours onto stainless steel alloy 316L implants. The implants are then irrigated using povidoneiodine solution with varying concentrations (5 and 10%) and durations (30, 60, and 180 seconds). The anti-biofilm effect was evaluated using plating and SEM methods to confirm its effectiveness. The process is repeated after 24 hours of postirrigation reincubation to detect any rebound growth. Results: No biofilm seen after irrigation with povidoneiodine at 5% and 10% concentrations at 30, 60 and 180 seconds, respectively, in both CFU count and SEM. This result is replicated after 24 hours of reincubation, in assessing for rebound growth. Conclusion: Our study supports that a minimum of 5% povidone-iodine with a minimum irrigation time of 30 seconds are effective at eliminating S. aureus biofilm on stainless steel alloy implants. Both CFU count and SEM yield similar value in validating the presence of biofilm. Additionally, SEM allows visualisation of the morphology of the biofilm.