Home › Cell Biology › Primary Cell Culture of Purified GABAergic or Glutamatergic Neurons Established through Fluorescence-activated Cell Sorting
Cell Biology JoVE (Open Access) Citable · DOI

Primary Cell Culture of Purified GABAergic or Glutamatergic Neurons Established through Fluorescence-activated Cell Sorting

DOI: 10.3791/58974-v
What you'll learn
  • ✓ Prepare and coat glass coverslips for neuronal culture
  • ✓ Dissociate hippocampal and cortical tissue from postnatal rodents
  • ✓ Sort and purify GABAergic or glutamatergic neurons using fluorescence-activated cell sorting
  • ✓ Culture purified neuronal populations in vitro
Protocol

Biopharma Insights This protocol describes a cell sorting based method for the purification and culture of fluorescent GABAergic or glutamatergic neurons from the neocortex and hippocampus of postnatal mice or rats.

Difficulty
advanced
Total time
~3–4 days (tissue dissociation ~2 hrs, cell sorting ~2–3 hrs, culture establishment ~2–3 days)
Model organism
Mouse or rat (postnatal, neocortex and hippocampus)
Biosafety
BSL-1

Steps

1
Coat glass coverslips with substrate

Prepare glass coverslips by coating with poly-D-lysine or laminin to promote neuronal cell adhesion and growth.

▶ 00:47
2
Dissociate hippocampal and cortical tissue

Extract and enzymatically dissociate postnatal mouse or rat hippocampus and neocortex to generate single-cell suspension.

▶ 02:22
3
Sort and culture purified neuronal populations

Use fluorescence-activated cell sorting (FACS) to isolate GABAergic or glutamatergic neurons and establish primary cultures on prepared coverslips.

▶ 07:13
4
Document cell culture purity and viability

Assess morphology, immunofluorescence staining, and viability of sorted neuronal populations to confirm successful purification and culture establishment.

▶ 09:19
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