In vitro synthesis of IgG directed against HIV components was detected by ELISA and Western blot assay of lymphocyte culture supernatants. Lymphocytes from HIV-infected individuals spontaneously produced antibody against HIV proteins very early in culture, suggesting in vivo activation of HIV-specific antibody-forming cells. The frequency of circulating B cells spontaneously secreting HIV-specific IgG was very high in some cases, but spontaneous HIV-specific antibody synthesis was not accompanied by polyclonal reactivation of B-cell clones of different specificity. The pattern of specificity of the anti-HIV antibody produced in vitro did not reflect the serum pattern consistently. These findings indicate a new approach potentially useful for the study of the immunobiology of HIV infection. The possible implications of the in vitro production of HIV-specific antibody for the diagnosis, prognosis and clinical management of this infection are also discussed.