expression seems to be negatively correlated with time in culture, being highest in freshly isolated cells and lowest in maintained primary cells and transformed lines. Most of these genes are in fact expressed at higher levels in pMGLs than in primary human fetal microglia (Fig. 5A, supplementary Fig. 2), supporting their identity as in vitro generated microglia. Notably, these genes are expressed at low levels in cultured macrophage cell lines as well as in previously described mouse ES-derived microglia 33. Conversely, genes such SLPI, SAA1/2, PRG4, CFP, CD5l, CRIP1 are expected to be highly expressed in peripheral macrophages, and low in microglia 11: these genes are in the lower expression quartile of our dataset. Genes such as GPR56 and BIN1 are expressed in microglia and in other CNS cell types, but have not been described in peripheral macrophages. Supplementary Figure 2c shows their expression in pMGLs, fMG, as well as differentiated NPCs. We performed unbiased hierarchical clustering of our data against previously published sequencing for primary and induced macrophages. As expected, all myeloid cells cluster away from parental iPS cells and NPCs. Strikingly, we find that pMGLs and primary fetal microglia cluster together, separately from other macrophages (Suppl. Figure 3b).
