Nitric oxide (·NO) has been implicated in immunopathogenesis of HIV-1 infection. Initial reports using low sensitive techniques showed elevated levels of ·NO in sera and tissues from seropositive patients. These results were not further supported using similar experimental approaches. To gain insight on ·NO deregulation during HIV-1 infection, we used recently described fluorescent probes with enhanced sensitivity to assess ·NO levels combined with iNOS mRNA expression in peripheral blood mononuclear cells (PBMC) from HIV-infected patients or after in vitro HIV-1 infection of normal cells. We demonstrate that PBMC from HIV-infected patients display a significant decrease of ·NO production and iNOS mRNA expression. Results from in vitro infection showed that HIV-1 induces a significant decrease in ·NO production and iNOS mRNA expression. Since ·NO could play a role in some key processes like apoptosis, regulation of immune responses and viral replication, these results could help in elucidating HIV-1 immunopathogenesis.