The MIT Center for Human Tissue Models of Infectious Diseases (MIT.HTMID) will study viral infections of five types of neural cells that are differentiated from human embryonic stem (ES) cells or human induced pluripotent stem (iPS) cells. In addition, MIT.HTMID research will address viral infections of human three dimensional (3D) cerebral organoids toward understanding Zika virus microcephaly. All cell work and tissue work for the Center’s research will be supported by the Human Cells and Tissues Core facility. The Core will be located at the Whitehead Institute, and will provide a continuous supply of cells for both HT.MID research projects. To prepare cells and organoids from iPS cells, human fibroblasts are reprogrammed into pluripotent iPS cells. Next, the pluripotent iPS cells are differentiated into multipotent neural progenitors (NPs). Neurons, astrocytes and oligodendrocyte are differentiated from NPs. To generate a homogenous population of NPs, a SMADinhibitionbased neural differentiation protocol is routinely used. Adherent cultures of control and mutant ESCs/iPSCs will be differentiated into neural rosettes expressing Pax6 and Nestin as detected by immunofluorescence, and further expanded as neuronal progenitors in the presence of bFGF. Multipotent NPs are differentiated into neurons. To generate neurons, NPs are cultured in a growth factor depleted medium that promotes terminal differentiation and maturation of a mixed populations of neurons. These neurons are electrophysiologically active, as measured on multielectrode arrays. Multipotent neural progenitors are separately differentiated into astrocytes or oligodendrocytes. To derive astrocytes from human NPs, we have developed protocols that allow the production of highly homogeneous populations. Specifically, FACSsorted PSANCAM/A2B5 glial progenitors, derived from NPs, are expanded in the presence of EGF and bFGF to + derive S100b+ immature astrocytes. Further maturation of this FACSsorted population, in the presence of CNTF, yields GFAP+ astrocytes. The cells can be sorted to high purity by magnetic cell sorting (MACS) against CD44 microbeads. To generate oligodendrocytes, A2B5+ glial progenitors will be expanded in bFGF and PDGFα, and further sorted by MACS against O4 and CD140a markers. These fated oligodendrocyte precursors are matured into MBP+ oligodendrocytes after growth factor removal and exposure to the thyroid hormone T3. The project investigators and core staff will meet regularly to plan work flow and anticipate cell needs for Projects 1 and 2. The Human Cells and Tissues core will be an integral and essential part of the MIT.HTMID Center, and its output will enhance the project’s potential for success