We describe the topographical distribution of microglial subpopulations during development of the human diencephalon and telencephalon. Brains from embryos and fetuses age 5–23.5 gestational weeks (gw) were subjected to single- and double-immunolabeling for lectin RCA-1 (Ricinus Communis Agglutinin 1), Iba1 (a microglial marker), CD68 (specific of macrophages), CD45 (marker for mononucleate cells of hematopoietic lineage), CD34 (expressed on endothelial cells), and MIB1 and Ki67 (markers for cell proliferation). At 5.5 gw the first intracerebral microglial cells were seen close to the meninges and choroid plexus near the di-telencephalic fissure. They were amoeboid and positive for Iba1, CD45, and RCA-1, whereas cells in the deep parenchyma expressed Iba1/CD68/RCA-1 and constituted clusters. In the developing diencephalon, microglial clusters were located in junctional regions of the white matter anlagen, most notably at the junctions of the internal capsule with the thalamic projections, the external capsule, and the cerebral peduncle. In the cortical anlagen, Iba1+/RCA-1/CD68+/CD45+ cells accumulated at 10–12 gw, constituting a tangential band at the junction between the cortical plate and the subplate. Between 10 and 16 gw microglial clusters increased markedly in size and cellular density. Contact between Iba1+ microglia and CD34+ blood vessels was clearly visible from 10–12 gw onward, first in microglial clusters of the white matter anlagen and subsequently throughout the parenchyma. From the middle of the second trimester onward microglial cells colonized the entire cerebral parenchyma, developed a ramified morphology, and downregulated their surface antigens, but remained more numerous in the white matter. J. Comp. Neurol. 499:565–582, 2006. © 2006 Wiley-Liss, Inc.