Astrocyte proliferation requires astrocyte elevated gene-1 (AEG-1). Astrocytes transfected with AEG-1 specific siRNA (siAEG-1) or non-targeting, scrambled siRNA (siCon) were immunostained with Ki67 or proliferating cell nuclear antigen (PCNA) (green, proliferation marker) and glial fibrillary acidic protein (GFAP) (red, astrocyte marker) antibodies. Untransfected and mock-transfected astrocytes were stained in parallel. Micrographs at 20 × original magnification of Ki67 or PCNA with GFAP costaining for non-transfected control (A1,A2), mock-transfected (B1,B2), siCon-transfected (C1,C2) and siAEG-1-transfected astrocytes (D1,D2) are shown. (E) This panel shows the number of Ki67 positive cells in ten micrographs from replicate wells for each condition (***P <0.001). (F) This panel shows the number of PCNA positive cells in ten micrographs from replicate wells for each condition (***P <0.001). All micrographs are 20 × original magnification, unless otherwise noted. Immunostaining data is representative of three individual donors assayed in triplicate.