We characterized the localization, phenotype, and some functions of plasmacytoid dendritic cells (pDCs) in the human spleen. pDCs were localized in the marginal zone and the periarteriolar region. Some were also found in the red pulp. pDCs were immature by phenotypic labeling, consistently with their capacity to internalize Dextran in a functional assay. In spleens from HIV-infected patients with thrombocytopenic purpura, these characteristics were unaffected. However, an accumulation of pDCs, but not myeloid dendritic cells (mDCs), was observed in some HIV+ patients, correlating with high proviral loads. Moreover, although undetectable in most HIV- patients, interferon-alpha (IFN-alpha) production was evidenced in situ and by flow cytometry in most HIV+ patients. IFN-alpha was located in the marginal zone. Surprisingly, IFN-alpha colocalized only with few pDCs, but rather with other cells, including T and B lymphocytes, mDCs, and macrophages. Therefore, pDCs accumulated in spleens from HIV+ patients with high proviral loads, but they did not seem to be the main IFN-alpha producers.