Enrichment and isolation of neurons from adult mouse brain for ex vivo analysis.
J Neurosci Methods
; 283: 15-22, 2017 May 01.
Article
en En
| MEDLINE
| ID: mdl-28336359
BACKGROUND: Isolation of neurons from the adult mouse CNS is important in order to study their gene expression during development or the course of different diseases. NEW METHODS: Here we present two different methods for the enrichment or isolation of neurons from adult mouse CNS. These methods: are either based on flow cytometry sorting of eYFP expressing neurons, or by depletion of non-neuronal cells by sorting with magnetic-beads. RESULTS: Enrichment by FACS sorting of eYFP positive neurons results in a population of 62.4% NeuN positive living neurons. qPCR data shows a 3-5fold upregulation of neuronal markers. The isolation of neurons based on depletion of non-neuronal cells using the Miltenyi Neuron Isolation Kit, reaches a purity of up to 86.5%. qPCR data of these isolated neurons shows an increase in neuronal markers and an absence of glial markers, proving pure neuronal RNA isolation. COMPARISON WITH EXISTING METHODS: Former data related to neuronal gene expression are mainly based on histology, which does not allow for high-throughput transcriptome analysis to examine differential gene expression. CONCLUSION: These protocols can be used to study cell type specific gene expression of neurons to unravel their function in the process of damage to the CNS.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Encéfalo
/
Separación Celular
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Técnicas de Cultivo Celular por Lotes
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Citometría de Flujo
/
Neuronas
Límite:
Animals
Idioma:
En
Revista:
J Neurosci Methods
Año:
2017
Tipo del documento:
Article
País de afiliación:
Alemania
Pais de publicación:
Países Bajos