A kind of naked mole rat cortical neuron culture method
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A technology of naked mole rat cortex and culture method, which is applied in the field of cell biology, can solve problems such as inapplicability to naked mole rats, and achieve good functional status and vitality, simple operation methods, and high reproducibility
Active Publication Date: 2019-07-02
SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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[0009] However, in the prior art, the methods for isolating, purifying and culturing cortical neurons of mice and rats are not suitable for naked mole rats. At present, there is no relevant report on the methods of isolating, purifying and culturing cortical neurons of naked mole rats in domestic and foreign literature.
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Embodiment 1
[0041] Example 1: Isolation, purification and culture of naked mole rat cortical neurons
[0042] 1. Experimental materials
[0043] Clean-grade naked mole rats at 65-70 days of pregnancy were provided by the Experimental Animal Center of the Second Military Medical University of the Chinese People's Liberation Army.
[0044] DNase was purchased from Solarbio Biotechnology Co., Ltd., NGF, 5-fluorouracil, trypsin, L-polylysine, penicillin and streptomycin mixture were purchased from Sigma Company, DMEM, low-sugar DMEM, fetal bovine serum from Australia, Neurobasal Culture medium, B27 serum-free additive factors, etc. were purchased from Thermo Fisher Scientific, and culture dishes, T25 and T75 culture flasks were purchased from Corning Company.
[0045] The mixed nerve cell culture medium consists of low-sugar DMEM medium containing 10% fetal bovine serum by volume.
[0046] Cortical neuron purification medium is Neurobasal + volume fraction 2% B27 + final concentration 10 μM...
Embodiment 2
[0055] Example 2: Identification of Naked Mole Rat Cortical Neurons
[0056] Identification of naked mole rat cortical neurons obtained in Example 1: 4% paraformaldehyde was used to fix the cortical neurons in serum-free medium, and combined with morphological identification, immunorefinement chemistry and other methods for identification.
[0057] 1. Cell morphology identification:
[0058] Identification methods described in references (Xuejun Chai, Shanting Zhao, Li Fan, Wei Zhang, XiLu, Hong Shao, Shaobo Wang, Lingzhen Song, Antonio Virgilio Failla, Bernd Zobiak, Hans G.Mannherz, Michael Frotscher, Reelin and cofilin cooperate during the migration of cortical neurons: a quantitative morphological analysis, Development, 2016, 143:1029-1040) the results are as follows figure 1 As shown, the naked mole-rat cortex neuron has a large cell body with one or several short dendrites and a long and thin axon at the other end.
[0059] 2. Immunocytochemical identification:
[0060...
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Abstract
The invention relates to the technical field of cell biology and in particular relates to a separation, purification and culture method of naked mole rat cortical neurons. The cortical neurons are separated from cerebral cortices of newly-born naked mole rats and are purified and cultured by comprehensively utilizing a plurality of types of culture mediums, and a reasonable culture method applicable to poikilothermal rodent mammal naked mole rat cortical neurons is explored. The method provided by the invention can be used for simply, conveniently, efficiently and economically obtaining a lot of naked mole rat cortical neurons with normal functional activity, and cells can still keep biological characteristics under an in-vivo state in an in-vitro environment through culture under a low-oxygen condition, so that special physiological functions of the naked mole rat cortical neurons can be conveniently and directly researched in a pure in-vitro cell culture model, and furthermore, an important theoretical basis is provided for exploring a biological mechanism and applying the biological mechanism to clinically relative fields.
Description
[0001] Technical field: [0002] The invention relates to the technical field of cell biology, in particular to a method for separating, purifying and culturing cells, in particular to a method for separating, purifying and culturing naked mole rat cortical neurons. [0003] Background technique: [0004] Although neurons do not have the highest proportion in the mammalian central nervous system, they are the most functionally important type of cells. Neurons play a vital role in synapse formation, synaptic signal transmission, and synaptic plasticity . The neurons in the cerebral cortex play the role of the control center in many functions of the body, including vision, hearing, smell, language, movement, and body surface sensation. [0005] The naked mole rat is a temperature-changing mammal. It can maintain its brain structure and function intact and carry out normal life activities in an unventilated cave with an oxygen concentration of only about 6%. Naked mole rats live...
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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/0793
CPCC12N5/0619C12N2500/40C12N2501/13
Inventor 崔淑芳杨文静孙伟汤球肖邦赵善民丛薇程继帅
Owner SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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