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An integrated cell isolation and purification method for rat dorsal root ganglion neurons
OBJECTIVE: Neurobiology studies are increasingly focused on the dorsal root ganglion (DRG), which plays an important role in neuropathic pain. Existing DRG neuron primary culture methods have considerable limitations, including challenging cell isolation and poor cell yield, which cause difficulty i...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
SAGE Publications
2019
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6683899/ https://www.ncbi.nlm.nih.gov/pubmed/31213102 http://dx.doi.org/10.1177/0300060519855585 |
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author | Shen, Huaishuang Gan, Minfeng Yang, Huilin Zou, Jun |
author_facet | Shen, Huaishuang Gan, Minfeng Yang, Huilin Zou, Jun |
author_sort | Shen, Huaishuang |
collection | PubMed |
description | OBJECTIVE: Neurobiology studies are increasingly focused on the dorsal root ganglion (DRG), which plays an important role in neuropathic pain. Existing DRG neuron primary culture methods have considerable limitations, including challenging cell isolation and poor cell yield, which cause difficulty in signaling pathway studies. The present study aimed to establish an integrated primary culture method for DRG neurons. METHODS: DRGs were obtained from fetal rats by microdissection, and then dissociated with trypsin. The dissociated neurons were treated with 5-fluorouracil to promote growth of neurons from the isolated cells. Then, reverse transcription polymerase chain reaction and immunofluorescence assays were used to identify and purify DRG neurons. RESULTS: Isolated DRGs were successfully dissociated and showed robust growth as individual DRG neurons in neurobasal medium. Both mRNA and protein assays confirmed that DRG neurons expressed neurofilament-200 and neuron-specific enolase. CONCLUSIONS: Highly purified, stable DRG neurons could be easily harvested and grown for extended periods by using this integrated cell isolation and purification method, which may help to elucidate the mechanisms underlying neuropathic pain. |
format | Online Article Text |
id | pubmed-6683899 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2019 |
publisher | SAGE Publications |
record_format | MEDLINE/PubMed |
spelling | pubmed-66838992019-08-19 An integrated cell isolation and purification method for rat dorsal root ganglion neurons Shen, Huaishuang Gan, Minfeng Yang, Huilin Zou, Jun J Int Med Res Pre-Clinical Research Reports OBJECTIVE: Neurobiology studies are increasingly focused on the dorsal root ganglion (DRG), which plays an important role in neuropathic pain. Existing DRG neuron primary culture methods have considerable limitations, including challenging cell isolation and poor cell yield, which cause difficulty in signaling pathway studies. The present study aimed to establish an integrated primary culture method for DRG neurons. METHODS: DRGs were obtained from fetal rats by microdissection, and then dissociated with trypsin. The dissociated neurons were treated with 5-fluorouracil to promote growth of neurons from the isolated cells. Then, reverse transcription polymerase chain reaction and immunofluorescence assays were used to identify and purify DRG neurons. RESULTS: Isolated DRGs were successfully dissociated and showed robust growth as individual DRG neurons in neurobasal medium. Both mRNA and protein assays confirmed that DRG neurons expressed neurofilament-200 and neuron-specific enolase. CONCLUSIONS: Highly purified, stable DRG neurons could be easily harvested and grown for extended periods by using this integrated cell isolation and purification method, which may help to elucidate the mechanisms underlying neuropathic pain. SAGE Publications 2019-06-19 2019-07 /pmc/articles/PMC6683899/ /pubmed/31213102 http://dx.doi.org/10.1177/0300060519855585 Text en © The Author(s) 2019 http://creativecommons.org/licenses/by-nc/4.0/ Creative Commons Non Commercial CC BY-NC: This article is distributed under the terms of the Creative Commons Attribution-NonCommercial 4.0 License (http://www.creativecommons.org/licenses/by-nc/4.0/) which permits non-commercial use, reproduction and distribution of the work without further permission provided the original work is attributed as specified on the SAGE and Open Access pages (https://us.sagepub.com/en-us/nam/open-access-at-sage). |
spellingShingle | Pre-Clinical Research Reports Shen, Huaishuang Gan, Minfeng Yang, Huilin Zou, Jun An integrated cell isolation and purification method for rat dorsal root ganglion neurons |
title | An integrated cell isolation and purification method for rat dorsal root ganglion neurons |
title_full | An integrated cell isolation and purification method for rat dorsal root ganglion neurons |
title_fullStr | An integrated cell isolation and purification method for rat dorsal root ganglion neurons |
title_full_unstemmed | An integrated cell isolation and purification method for rat dorsal root ganglion neurons |
title_short | An integrated cell isolation and purification method for rat dorsal root ganglion neurons |
title_sort | integrated cell isolation and purification method for rat dorsal root ganglion neurons |
topic | Pre-Clinical Research Reports |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6683899/ https://www.ncbi.nlm.nih.gov/pubmed/31213102 http://dx.doi.org/10.1177/0300060519855585 |
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