Cell lines originally derived from malignant tumours of the brain were infected by diverse human immunodeficiency virus types 1 and 2 (HIV-1 and HIV-2) isolates. By surface immunofluorescence it was shown that susceptible cells did not bear the CD4 antigen. They were also non-permissive for the formation of plaques by vesicular stomatitis virus pseudotypes and did not form syncytia with HIV-producing cells. Virus production was of low titre, and reverse transcriptase and the p24 antigen were consistently undetectable in the culture supernatants. Output virus could be detected by cocultivation with a sensitive T cell line, C8166, by the culture of supernatant medium with T cells and by detection of proviral HIV DNA after amplification. A higher multiplicity of input virus was required to establish a brain cell infection than was required for T lymphocytes or monocytes. Some HIV-susceptible brain cells contained mRNA for CD4 but infection was not blocked by anti-CD4 antibodies. Apparently HIV infection of these cells does not involve CD4 as the cellular receptor.