B cell dysfunction is a well-studied complication of HIV infection in adults. Data on B cell differentiation in normal and HIV-infected children are lacking. We show the distribution of B cell subsets and immunoglobulin levels in HIV-infected children compared with controls. Furthermore, we observe the long-term B cell reconstitution of vaccine-specific immunity after antiretroviral therapy (ART). Phenotype of B cells (naive, non-switched memory, switched memory) was analyzed in 48 infected children and 62 controls. In nine HIV-infected children, functional reconstitution was quantified by tetanus-specific antibodies and by performing a lymphocyte transformation test (LTT) in a longitudinal approach. Switched memory B cells are significantly reduced in HIV-infected children. Vaccine-specific antibodies and response to LTT increase after initiation of ART. Our data indicate a significant dysfunction in the B cell system, despite effective ART. Partial reconstitution of humoral immunity may have therapeutic implications in a subset of HIV-infected children.