Hi
I've been trying to isolate E15 spinal cord neurons. After isolation and digestion, I incubate the cells with Neural Base Media and 2% B27.
Initially I used to add FBS to the media. Because of over growth of fibroblasts I stopped adding FBS, which stopped fibroblasts from growing. After 7-8 days I see the neural cells with some extensions but it seems the cell extensions are not growing fast enough and after a while they die. What do I need to change?
thanks
mostafa
Remove disgestion, use technique by passing tissue through pipett opening, which you diminish several times (warm up pipett and pull, let cool the glass!) Collect spinal cords in chemically defined medium, mechanically dissociation using the Pasteur pipett methode of Romijn et al., 1984. Omitt antibiotics. Add nerve growth factor to final conc. 100 ul/ml (Marani, 1988). see Marani et al., 1988 Neuroendocrinology 48: 445-452; Romijn et al., Neurosci and Bio behav Rev 8: 301-344.
Succes, Marani
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