Background: Out-of-pocket costs for opioid agonist treatment (OAT) constitute a barrier to treatment entry and retention.This study examines OAT clients’ total out-of-pocket costs (including dispensing fees, travel costs and OAT-related appointment costs) in different treatment settings (public clinics, community pharmacies, and private clinics). Methods: Cross-sectional survey of 402 people with opioid drug use (OUD) in New South Wales (NSW), Victoria (VIC), Tasmania (TAS), Australia; 266 clients (66%) currently receiving methadone, buprenorphine or buprenorphine-naloxone treatment were asked about dispensing fees, travel costs and OAT-related appointment costs in the past 28 days. A two-part regression model was used to deal with non-normal distributions of costing data (right skew and excess zeros). Results: Among clients currently receiving OAT, 87% paid out-of-pocket costs. Among those who paid out-of-pocket costs (N=194), travel costs accounted for more than half of total costs (52%), followed by dispensing fees (44%). The mean monthly total out-of-pocket costs were AU$135 (SD: AU$121) for public clinics, AU$161 (SD: AU$110) to AU$214 (SD: AU$166) for community pharmacies and AU$355 (SD: AU$159) for private clinics. Compared to participants in NSW private clinics, those at public clinics paid one third the total out-of-pocket costs (coefficient = 0.33; 95%CI = 0.23-0.48) and those at NSW, TAS, VIC pharmacies paid approximately half the costs (coefficient = 0.58; 95%CI = 0.42-0.79; coefficient = 0.51; 95%CI = 0.36-0.72; coefficient = 0.47; 95%CI = 0.34-0.66, respectively). People in OAT for more than a year paid half the total out-of-pocket costs, compared with those in OAT less than a year (coefficient = 0.49, 95%CI = 0.31-0.77). Conclusions: Participants in the current study spent one-eighth of their income on out-of-pocket costs associated with OAT representing a substantial financial burden. Total out-of-pocket costs disproportionately affects those who are newer in treatment and receiving fewer unsupervised doses. Considering and addressing total out-of-pocket costs, especially travel costs and dispensing fees, to clients is critical to prevent cost from being a barrier from receiving effective care.