Figure 2.
Identification of transfected cells by immunoelectromicroscopy. A, GFP-encoding Mokola vector was injected into the RMS elbow of Ara-C treated mice. Animals were killed for analysis at 1 d.p.i. B, D, GFP-expressing cells (B1, B2) showing light cytoplasm, irregular contours of cell surface and nonclumped chromatin typical for type-B1 astrocytes (B) or clumped chromatin typical for candidate stem cell type-B2 astrocytes (D). C, E, Details of panels B and D, respectively (white squares) showing examples of immunostaining of GFP in astrocyte cytoplasm. The electron dense precipitate of oxidized DAB was intentionally maintained at low intensity to facilitate ultrastructural identification of B astrocytes (arrows). F, GFP-expressing cell showing intentionally enhanced immunostaining of GFP in astrocytic cytoplasm. G, GFP-expressing cell showing typical astrocytic end foot apposed to the abluminal face of a blood vessel (arrow). H, Panorama showing the ultrastructure of the RMS elbow 1 d after the end of Ara-C treatment (Ara-C). Chains of neuroblasts were not observed and only astrocytes (B) could be detected. A mitotic figure is also shown (M). I, Panorama showing the ultrastructure of the RMS elbow in nontreated animals (Control) where chains of migrating neuroblasts (A) interspersed with astrocyte-like glial cells (B) are clearly distinguished. Magnification 1200× (B, D, F), 5000× (C, E), 2000× (G), 800× (H, I). Scale bars: 5 μm (B, D, F-I) and 1 μm (C, E). IEM, immunoelectron-microscopy; SVZ, subventricular zone; RMS, rostral migratory steam; OB, olfactory bulb; LV, lateral ventricle; BV, blood vessel.