PMID- 28518109
OWN - NLM
STAT- MEDLINE
DCOM- 20180119
LR  - 20190510
IS  - 1940-087X (Electronic)
IS  - 1940-087X (Linking)
IP  - 123
DP  - 2017 May 10
TI  - Assessment of Neuronal Viability Using Fluorescein Diacetate-Propidium Iodide 
      Double Staining in Cerebellar Granule Neuron Culture.
LID - 10.3791/55442 [doi]
LID - 55442
AB  - Primary cultured Cerebellar Granule Neurons (CGNs) have been widely used as an in 
      vitro model in neuroscience and neuropharmacology research. However, the 
      co-existence of glial cells and neurons in CGN culture might lead to biases in 
      the accurate assessment of neuronal viability. Fluorescein diacetate (FDA) and 
      Propidium Iodide (PI) double staining has been used to measure cell viability by 
      simultaneously evaluating the viable and dead cells. We used FDA-PI double 
      staining to improve the sensitivities of the colorimetric assays and to evaluate 
      neuronal viability in CGNs. Furthermore, we added blue fluorescent DNA stains 
      (e.g., Hoechst) to improve the accuracy. This protocol describes how to improve 
      the accuracy of assessment of neuronal viability by using these methods in CGN 
      culture. Using this protocol, the number of glial cells can be excluded by using 
      fluorescence microscopy. A similar strategy can be applied to distinguish the 
      unwanted glial cells from neurons in various mixed cell cultures, such as primary 
      cortical culture and hippocampal culture.
FAU - Jiajia, Lin
AU  - Jiajia L
AD  - Ningbo Key Laboratory of Behavioral Neuroscience, Zhejiang Provincial Key 
      Laboratory of Pathophysiology, School of Medicine, Ningbo University.
FAU - Shinghung, Mak
AU  - Shinghung M
AD  - Department of Applied Biology and Chemistry Technology, Institute of Modern 
      Chinese Medicine, The Hong Kong Polytechnic University.
FAU - Jiacheng, Zheng
AU  - Jiacheng Z
AD  - Ningbo Key Laboratory of Behavioral Neuroscience, Zhejiang Provincial Key 
      Laboratory of Pathophysiology, School of Medicine, Ningbo University.
FAU - Jialing, Wang
AU  - Jialing W
AD  - Ningbo Key Laboratory of Behavioral Neuroscience, Zhejiang Provincial Key 
      Laboratory of Pathophysiology, School of Medicine, Ningbo University.
FAU - Dilin, Xu
AU  - Dilin X
AD  - Ningbo Key Laboratory of Behavioral Neuroscience, Zhejiang Provincial Key 
      Laboratory of Pathophysiology, School of Medicine, Ningbo University.
FAU - Shengquan, Hu
AU  - Shengquan H
AD  - Department of Applied Biology and Chemistry Technology, Institute of Modern 
      Chinese Medicine, The Hong Kong Polytechnic University.
FAU - Zaijun, Zhang
AU  - Zaijun Z
AD  - Institute of New Drug Research, Guangdong Provincial Key Laboratory of 
      Pharmacodynamic, Constituents of Traditional Chinese Medicine & New Drug 
      Research, College of Pharmacy, Jinan University.
FAU - Qinwen, Wang
AU  - Qinwen W
AD  - Ningbo Key Laboratory of Behavioral Neuroscience, Zhejiang Provincial Key 
      Laboratory of Pathophysiology, School of Medicine, Ningbo University.
FAU - Yifan, Han
AU  - Yifan H
AD  - Department of Applied Biology and Chemistry Technology, Institute of Modern 
      Chinese Medicine, The Hong Kong Polytechnic University; International Joint 
      Laboratory (SYSU-PolyU HK) of Novel Anti-Dementia Drugs of Guangdong.
FAU - Wei, Cui
AU  - Wei C
AD  - Ningbo Key Laboratory of Behavioral Neuroscience, Zhejiang Provincial Key 
      Laboratory of Pathophysiology, School of Medicine, Ningbo University; Department 
      of Applied Biology and Chemistry Technology, Institute of Modern Chinese 
      Medicine, The Hong Kong Polytechnic University; chester0151005@163.com.
LA  - eng
PT  - Journal Article
PT  - Video-Audio Media
DEP - 20170510
PL  - United States
TA  - J Vis Exp
JT  - Journal of visualized experiments : JoVE
JID - 101313252
RN  - 0 (Coloring Agents)
RN  - 0 (Culture Media)
RN  - 0 (Fluoresceins)
RN  - 0 (Fluorescent Dyes)
RN  - 36015-30-2 (Propidium)
RN  - YL39R93PRE (diacetylfluorescein)
SB  - IM
MH  - Animals
MH  - *Cell Survival
MH  - Cells, Cultured
MH  - Cerebellum/*cytology
MH  - Colorimetry
MH  - *Coloring Agents
MH  - Culture Media
MH  - Cytoplasmic Granules
MH  - *Fluoresceins
MH  - *Fluorescent Dyes
MH  - Microscopy, Fluorescence
MH  - Neuroglia
MH  - Neurons/*physiology
MH  - *Propidium
MH  - Rats
MH  - Rats, Sprague-Dawley
MH  - Reproducibility of Results
MH  - Staining and Labeling
PMC - PMC5607932
EDAT- 2017/05/19 06:00
MHDA- 2018/01/20 06:00
PMCR- 2019/05/10
CRDT- 2017/05/19 06:00
PHST- 2017/05/19 06:00 [entrez]
PHST- 2017/05/19 06:00 [pubmed]
PHST- 2018/01/20 06:00 [medline]
PHST- 2019/05/10 00:00 [pmc-release]
AID - 55442 [pii]
AID - 10.3791/55442 [doi]
PST - epublish
SO  - J Vis Exp. 2017 May 10;(123):55442. doi: 10.3791/55442.