Isolating Sensory Neurons and Co-Culturing with Natural Killer Cells
Transcription
Take a dorsal root ganglion or DRG, a cluster of sensory neurons, in a cold, serum-containing medium to protect the neurons during subsequent processing.
Centrifuge, then remove the supernatant.
Add a phosphate buffer containing digestive enzymes and incubate with mild agitation.
The enzymes dissociate the extracellular matrix, or ECM, and DRG tissue, loosening the cells.
Add a serum-containing medium to inactivate the enzymes.
Centrifuge, then discard the supernatant.
Resuspend the DRG and mechanically dissociate the tissue using pipettes of progressively smaller sizes to obtain a cell suspension.
Layer the cells onto a density gradient of bovine serum albumin.
Centrifuge. The neurons settle to the bottom, forming a pellet.
Discard the layers containing tissue debris.
Resuspend the neurons in a culture medium.
Add the neurons to an ECM-coated culture dish and incubate.
Remove the medium and add natural killer or NK cells.
Add a medium to co-culture and study the interaction between these cells.
