We propose to determine the contribution of covalently closed circular DNA (cccDNA) and host genome (iDNA) to hepatitis B surface antigen (HBsAg) at the single hepatocyte level before and during nucleos(t)ide (NUC) therapy in 5 HIV-HBV co-infected individuals with paired archived liver tissue. Our research will broadly impact the field by using novel techniques to determine the proportions of HBsAg from cccDNA or iDNA, which will inform the rational design of therapies for HBV cure.