Fourteen isolates of Lassa virus were recovered in Vero cell cultures from material—serum, pleural fluid, urine, and throat washings—of four cases of Lassa fever. Viremia of 1 to 2 weeks' duration, with TCD50 titers ranging from 2 to 4.5 dex per ml, was observed. The agent did not infect the Aedes aegypti and Aedes albopictus continuous cell lines. When newborn and adult mice were inoculated intracerebrally with Lassa virus, complement-fixing and neutralizing antibodies were detected in their serum; in addition, adult mice showed signs closely resembling those seen in adult mice inoculated with lymphocytic choriomeningitis (LCM) virus. Lassa virus was isolated from urine of infected mice as late as 83 days after inoculation. Multiplication of Lassa virus in Vero cell cultures was not inhibited by the incorporation of 5-bromodeoxyuridine in the medium; hence the virus probably contains ribonucleic acid. The finding that the agent is susceptible to the action of sodium deoxycholate suggests the presence of a lipid-containing envelope. Electronmicroscopy studies reveal a spherical shape. Filtration studies indicate a diameter of the virus between 70 and 150 mµ. The 14 isolates, insofar as studied, are indistinguishable from one another. In extensive serologic studies, Lassa virus has been compared with and found distinct from numerous arboviruses and other viruses. By complement-fixation test, it cross-reacts to a low degree with LCM virus, and possibly also with some members of the Tacaribe group.