The dendritic cell (DC)-specific molecule DC-SIGN is a receptor for the HIV-1 envelope glycoprotein gp120 and is essential for the dissemination of HIV-1. DC-SIGN is expressed by DCs, both monocyte-derived DCs and DCs in several tissues, including mucosa and lymph nodes. To identify a DC-SIGN(+) DC in blood that may be involved in HIV-1 infection through blood, we have analyzed the expression of DC-SIGN in human blood cells. Here we describe the characterization of a subset of DCs in human blood, isolated from T-/NK-/B-cell-depleted peripheral blood mononuclear cells (PBMCs) on the basis of expression of DC-SIGN. This subset coexpresses CD14, CD16, and CD33 and is thus of myeloid origin. In contrast to CD14(+) monocytes, DC-SIGN(+) blood cells display a DC-like morphology and express markers of antigen-presenting cells, including CD1c, CD11b, CD11c, CD86, and high levels of major histocompatibility complex (MHC) class I and II molecules. This DC population differs from other described CD14(-) blood DC subsets. Functionally, DC-SIGN(+) blood DCs are able to stimulate proliferation of allogeneic T cells and can produce tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) upon activation with lipopolysaccharide (LPS). When they encounter HIV-1, low amounts of these blood DC-SIGN(+) DCs enhance infection of T lymphocytes in trans, whereas blood monocytes and CD14(-) blood DCs are not capable of transmitting HIV-1. Therefore DC-SIGN(+) blood DCs can be the first target for HIV-1 upon transmission via blood; they can capture minute amounts of HIV-1 through DC-SIGN and transfer HIV-1 to infect target T cells in trans.