The effect of human serum with or without Epstein-Barr virus (EBV) antibodies was characterized on virus production in P3HR-1 cells. Cell culturing with EBV-seropositive sera reduced both production of infectious virus and amounts of virion DNA in the supernat-ants. EBV DNA was also reduced in the cells. Such reductions in cell-associated EBV DNA depended upon the concentration of seropositive serum and incubation time. Decreased frequencies of productive EBV DNA-replicating cells were observed in cell populations which had reduced levels of cell-associated EBV DNA. The inhibitory effect of seropositive serum was reversed upon switching the cells to medium with seronegative serum. In serial sera of an acute infectious mononucleosis patient the EBV DNA-reducing activity arose in parallel to antibodies against EBV membrane antigen and nuclear antigen. Possible mechanisms were discussed for antibody-mediated inhibition of EBV production.