Figure 6

Confrontation assays illustrate astrocytes express calponin. Confrontation assays were set up between OECs/astrocytes (A, B) and Schwann cells/astrocytes (C, D). A) OECs mingled with astrocytes while B) Schwann cells formed a boundary with astrocytes. Astrocytes expressed GFAP (red) and OECs or Schwann cells expressed p75^NTR (green). B, D) When these cultures were labeled with calponin (green), GFAP (red) and DAPI for nuclei of the Schwann cells or OECs (blue) it could be seen that both OECs and Schwann cells lacked calponin expression (green immunoreactivity alone) but astrocytes expressed calponin (red and green immunoreactivity). E-G) Astrocytes generated from P1 cortex and P1 neurospheres were immunolabeled with antibodies to GFAP (red) and to calponin (green). Neonatal cortical astrocytes (CAs) were over 98% pure as assessed by GFAP expression (E) and expressed fibrillar calponin heterogeneously (E-F). Astrocytes generated from neurospheres (NsAs) by differentiation in DMEM- 10% FBS also expressed calponin heterogeneously in culture (G). H) No cell staining occurred in control cultures when the primary antibody was excluded (DAPI, blue). Images for Figures 6A-D, G were collected using the fluorescent microscope and images for Figures 6E, F, H were collected using confocal microscopy.