A kind of naked mole rat drg neuron culture method
A culture method and neuron technology, applied in the field of cell biology, can solve problems such as inapplicability to naked mole rats, and achieve high reproducibility and simple operation methods
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Embodiment 1
[0040] Example 1: Isolation, purification and culture of naked mole rat DRG neurons
[0041] 1. Experimental materials
[0042] Clean-grade naked mole rats at 55-60 days of pregnancy were provided by the Experimental Animal Center of the Second Military Medical University of the Chinese People's Liberation Army.
[0043] DNase I was purchased from Solarbio Biotechnology Co., Ltd., NGF, 5-fluorouracil, trypsin, D-polylysine, collagen penicillin and streptomycin mixture, etc. were purchased from Sigma Company, DMEM, low-sugar DMEM, Australian source fetal Bovine serum, Neurobasal medium, and B27 serum-free additive factors were purchased from Thermo Fisher Scientific, and 24-well culture plates were purchased from Corning.
[0044] The components of the mixed nerve cell culture medium are low-sugar DMEM medium containing 10% fetal bovine serum by volume fraction, and the DRG neuron purification medium is Neurobasal+2% B27 by volume fraction+final concentration of 10 μM 5-fluoro...
Embodiment 2
[0053] Example 2: Identification of Naked Mole Rat DRG Neurons
[0054] Identification of naked mole-rat DRG neurons obtained in Example 1: DRG neurons in serum-free medium were fixed with 4% paraformaldehyde, and identified by morphological identification, immunorefinement chemistry and other methods.
[0055] 1. Cell morphology identification:
[0056] Identification methods described in references (Yang Y, Huang J, Mis MA, Estacion M, Macala L, ShahP, Schulman BR, Horton DB, Dib-Hajj SD, Waxman SG.Nav1.7-A1632G Mutation from aFamily with Inherited Erythromelalgia : Enhanced Firing of Dorsal Root Ganglia Neurons Evoked by Thermal Stimuli. J Neurosci. 2016 Jul 13; 36(28):7511-22.).
[0057] The result is as figure 1 (low magnification lens) and figure 2 (High magnification) shows that under the light microscope, the naked mole-rat DRG neurons have larger cell bodies, one or several shorter dendrites, and one elongated axon at the other end.
[0058] 2. Immunocytochemical...
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