We previously reported that infectivities of human and other non-human lentiviruses including human immunodeficiency virus type 1 (HIV-1) are activated by desialylation of the virion surface [H. Hu et al., J Virol 70: 7,462-7,470 (1996)]. The present study was designed to determine whether neuraminidase (NA) is useful for isolation of HIV-1 from patients. Peripheral blood mononuclear cells (PBMC) or CD4+ cells isolated from the PBMC of infected individuals were cocultured with PBMC or CD4+ cells from uninfected healthy donors, and the efficiency and frequency of virus isolation in the presence of NA were compared with those by a routine conventional procedure. In a total of 41 isolation trials from 28 individuals, the presence of NA markedly increased the frequency of isolation. Furthermore, both the day when virus was first detected and the day when the virus titer was the highest in the cultures were significantly earlier in the presence of NA than in the absence of NA. The deduced amino acid sequences of the V2 and V3 regions of gp120 were identical or very similar between the isolates obtained in the presence or absence of NA, suggesting that both isolation procedures selected a similar population. NA was thus found to facilitate HIV-1 isolation and its use is recommended particularly when isolation is negative by the conventional procedures.