Dissociating and Culturing Neurons from Hippocampal Tissue Samples
Transcription
Start with hippocampal tissue samples containing neurons and various non-neuronal cells.
Add a protease enzyme solution. The enzyme degrades the tissue’s extracellular matrix, initiating cell dissociation.
Wash with a plating medium to remove residual enzymes.
Pipette the digested tissue repeatedly to mechanically dissociate the cells from the tissue, forming a single-cell suspension.
Centrifuge and remove the supernatant. Resuspend the cells in the plating medium.
Seed the cells onto a poly-D-lysine-coated coverslip in a multi-well plate. Lysine enhances cell attachment.
Replace the plating medium with a neuron maintenance medium containing essential nutrients for neuron survival.
Refresh half of the media with maintenance media containing arabinosylcytosine, or Ara-C.
Ara-C enters the cells and selectively inhibits the growth of non-neuronal cells, leading to their death.
Replace the media with fresh maintenance media to support neuron survival.
