Astroglial cell morphogenesis in Matrigel

Matrigel (0.5 mL; Corning #354234) was applied to the bottom of a 35 mm tissue culture dish to harden for 30 min at 37°C. Trypsin-EDTA was used to remove the cells. The cells were then washed with growth medium 1x and then resuspended at 2 x 10⁵ cells/mL in growth medium not containing FBS. Then 2 x 10⁵ cells in 2 mL was added to the Matrigel-coated plates and incubation continued at 33⁰ C for 18 hours. To quantitatively assess the data, we determined the mean numbers of branches under 5 high-power fields (x100) using the following software for analysis http://image.bio.methods.free.fr/ImageJ/?Angiogenesis-Analyzer-for-ImageJ&artpage=3-6#outil_sommaire_3. Note increasing the incubation time did not enhance branching morphogenesis.