To obtain additional information about the early steps in the infectious cycle of vaccinia, agents were used which interfere with the normal uptake of virus particles and with the penetration of their DNA to intracellular sites of function. Treatment of host cells with sodium fluoride causes an almost complete but reversible arrest of virus uptake. Virus remaining thereby on the surface of cells can be rendered noninfective by antiserum, indicating that phagocytosis is an obligatory step for initiation of infection. Inactivation of vaccinia by heating or antibody, which abolishes infectivity, does not prevent uptake of virus into phagocytic vacuoles but does block the normal transfer of viral cores from the vacuoles. Instead, the trapped particles, including their DNA, are gradually but totally destroyed, probably by lytic enzymes within the vacuoles. The relationship of these findings to the normal orderly penetration and release of the viral genome is discussed.