To explore the spatiotemporal dynamics of human macrophage specification, we began with an unbiased approach by sorting all the immune cells (CD45+ cells) for scRNA-seq analysis from 19 organs/tissues of 52 human embryos and fetuses. In total, we collected 367 prenatal samples spanning 19 time points encompassing postconceptional weeks (PCW) 4–26, from the mid-embryonic stage (Carnegie stage 11, CS11) to the early third trimester (117 26 PCW). A total of 293,095 immune cells from the embryonic, fetal, and adult tissues (yolk sac, embryo, head, AGM, heart, liver, GI tract, limb, lung, kidney, adrenal gland, spinal cord, brain, male/female gonad, thymus, skin, bone marrow, and spleen) were profiled and passed the quality control.