BACKGROUND: The prevalence of HIV and Hepatitis C Virus (HCV) are significantly lower among people who inject drugs (PWID) in San Diego, CA, USA compared with PWID in Tijuana, Mexico, located directly across the border. We investigated associations between cross-border injection drug use (IDU), HIV and HCV seroprevalence and engagement in injecting risk behaviours while on each side of the border. METHODS: Using baseline interviews and serologic testing data from STAHR II, a longitudinal cohort study of PWID in San Diego, bivariate and multivariable logistic regression analyses examined associations between recent (past six months) cross-border IDU and HIV and HCV antibody seropositivity, socio-demographics, drug use characteristics, and participants' connections to, and perceptions about Mexico. Chi-squared tests and McNemar tests examined associations between cross-border IDU and injecting risk behaviours. RESULTS: Of the 567 participants (93% U.S.-born, 73% male, median age 45 years), 86 (15%) reported recent cross-border IDU. Cross-border IDU was not associated with HIV (OR: 0.85, 95% CI: 0.37-1.95) or HCV seropositivity (OR: 1.01, 95% CI: 0.62-1.65). Age, identifying as Hispanic or Latino/a, and being concerned about risk of violence when travelling to Mexico were independently associated with decreased odds of recent cross-border IDU. Injecting cocaine at least weekly, having ever lived in Mexico and knowing PWID who reside in Mexico were associated with increased odds of recent cross-border IDU. PWID who reported cross-border IDU were significantly less likely to engage in receptive needle sharing, equipment sharing, and public injection while in Mexico compared with in San Diego (all p<0.001). CONCLUSION: Prevalence of HIV and HCV infection was similar among PWID who had and had not injected in Mexico, possibly due to practising safer injecting while in Mexico. Research is needed to elucidate contextual factors enabling U.S. PWID to inject safely while in Mexico.