worthington 腦組織分離指南

發(fā)布者：艾美捷科技發(fā)布時間：2022-05-31

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原代細(xì)胞分離中，針對腦組織涉及到微血管內(nèi)皮、腦內(nèi)皮細(xì)胞、神經(jīng)元和小膠質(zhì)細(xì)胞、微血管等等的解離通常會使用酶，如何選擇合適的酶來完成對應(yīng)腦組織的解離，以及原代細(xì)胞采集的常規(guī)操作， 艾美捷針對腦相關(guān)的組織分類解離分種屬、不同的細(xì)胞、酶組合推薦，推薦濃度、培養(yǎng)基以及參考文獻(xiàn)，詳細(xì)匯總指南：腦組織分離指南

本技術(shù)指南描述了標(biāo)準(zhǔn)實(shí)驗(yàn)室程序；為建立細(xì)胞分離方案提供了一種合乎邏輯的實(shí)驗(yàn)方法；并列出了許多組織特異性參考資料，以幫助開發(fā)有效的方法。更多其他組織的解離指南，點(diǎn)擊查看：各組織分離指南。

腦
種屬	種屬詳情	細(xì)胞名稱	酶組合推薦，含推薦濃度	培養(yǎng)基	參考文獻(xiàn)
牛	Bovine	微血管內(nèi)皮 / Microvascular endothelial	Neutral Protease: 0.005%	Medium 199	Kanda, T., Yoshino, H., Ariga, T., Yamawaki, M., and Yu, R.: Glycosphingolipid Antigens in Cultured Microvasular Bovine Brain Endothelial Cells: Sulfoglucuronosyl Paragloboside as a Target of Monoclonal IgM in Demyelinative Neuropathy, J Cell Biol 126 (1), 235, 1994
牛	Bovine	腦內(nèi)皮細(xì)胞 / Brain endothelial cells	Collagenase Type 2: 0.35%	DMEM	Wolburg H, Neuhaus J, Kniesel U, Krauss B, Schmid EM, Ocalan M, Farrell C, Risau W: Modulation of tight junction structure in blood-brain barrier endothelial cells. Effects of tissue culture, second messengers and cocultured astrocytes, J Cell Sci 107, 1347, 1994
牛	Bovine	內(nèi)皮 / Endothelial	Neutral Protease: 0.125%	MEM	Miller, D., Audus, K., and Borchardt, R.: Application of Cultured Endothelial Cells of the Brain Microvasculature in the Study of the Blood-Brain Barrier, J Tiss Cul Meth 14, 217, 1992
牛	Bovine, adult	腦動脈，內(nèi)皮 / Cerebral artery，Endothelial	Collagenase: 0.2%	HBSS	Machi, T., Kassell, N.F., and Scheld, W.M.: Isolation and Characterization of Endothelial Cells From Bovine Cerebral Arteries, In Vitro Cell Dev Biol 26, 291, 1990
牛	Bovine	毛細(xì)血管內(nèi)皮 / Capillary endothelial	Collagenase: 0.1%	DMEM	Estrada, C., Bready, J., Berliner, J., and Cancilla, P.: Choline Uptake by Cerebral Capillary Endothelial Cells in Culture, J Neurochem 54, 1467, 1990
牛	Bovine	內(nèi)皮 / Endothelial	Neutral Protease: 0.5%	MEM	Audus, K., and Borchardt, R.: Characterization of an In Vitro Blood-Brain Barrier Model System for Studying Drug Transport and Metabolism, Pharm Res 3 (2), 81, 1986
牛	Bovine	內(nèi)皮，腦動脈 / Endothelial，Brain arteries	Collagenase Type 2: 0.2%	Dulbecco's PBS	Goetz, I.., Warren, J., Estrada, C., Roberts, E., and Krause, D.: Long-Term Cultivation of Arterial and Capillary Endothelium From Adult Bovine Brain, In Vitro Cell Dev Biol 21, 172, 1985
牛	Calf	少突膠質(zhì)細(xì)胞，神經(jīng) / Oligodendroglia，Neural	Trypsin: 0.1%	(see reference)	Poduslo, S., Miller, K., and McKhann, G.: Metabolic Properties of Maintained Oligodendroglia Purified from Brain, J Biol Chem 253, 1592, 1978
豚鼠	Guinea-pig, 200-400 g	神經(jīng)元 / Neurons	Trypsin: 0.06-0.08%	PIPES saline	Kay, A.R., and Wong, R.K.S.: Isolation of Neurons Suitable for Patch-Clamping from Adult Mammalian Central Nervous Systems, J Neurosci Methods 16, 227, 1986
人	Human	小膠質(zhì)細(xì)胞 / Microglia	Collagenase Type 1: 300 u/ml； Trypsin: 0.125%	DMEM	Mizee, M., Miedema, S., van der Poel, M., Adelia , S., van Strien, M., Melief, J., Smolders, J., Hendrickx, D., Heutinck, K., Hamann, J. and Huitinga, I.: Isolation of Primary Microglia from the Human Post-Mortem Brain: Effects of Ante- and Post-Mortem Variables., Acta Neuropathol 5, 16, 2017
人	Human, adult	神經(jīng)元 / Neuronal	Papain: 20 u/ml	Neurobasal	Spaethling, J., Na, Y., Lee, J., Ulyanova, A., Baltuch, G., Bell, T., Brem, S., Chen, H., Dueck, H., Fisher, S., Garcia, M., Khaladkar, M., Kung, D., Lucas, T., O'Rourke, D. and Stefanik, D.: Primary Cell Culture of Live Neurosurgically Resected Aged Adult Human Brain Cells and Single Cell Transcriptomics., Cell Rep 18, 791-803, 2017
人	Human, fetal and mature	星形膠質(zhì)細(xì)胞和神經(jīng)元 / Astrocytes and neurons	Papain: 7.5-20 u/ml	RPMI	Zhang, Y., Sloan, S., Clarke, L., Caneda, C., Plaza, C., Blumenthal, P., Vogel, H., Steinberg, G., Edwards, M., Li, G., Duncan, J., Cheshier, S., Shuer, L., Chang, E., Grant, G., Gephart, M. and Barres, B.: Purification and Characterization of Progenitor and Mature Human Astrocytes Reveals Transcriptional and Functional Differences with Mouse., Neuron 89, 37-53, 2016
人	Human	小膠質(zhì)細(xì)胞 / Microglia	Neutral Protease: 10 u/ml； Papain: 2.5 u/ml	Hibernate A	Rustenhoven, J., Park Thomas, I., Schweder, P., Scotter, J., Correia, J., Smith, A., Gibbons, H., Oldfield, R., Bergin, P., Mee, E., Faull, R., Curtis, M., Scott, G. and Dragunow, M.: Isolation of Highly Enriched Primary Human Microglia for Functional Studies., Sci Rep 6, 19371, 2016
人	Human, 25-81 yr	腦腫瘤 / Brain tumor	Neutral Protease: 0.11 u/ml； Collagenase Type 4: 0.05%； Hyaluronidase: 1,000 u/ml； Deoxyribonuclease I: 5 u/ml；	HBSS	Volovitz, I, Shapira, N., Ezer, H., Gafni, A., Lustgarten, M., Alter, T., Ben-Horin, I., Barzilai, O., Shahar, T., Kanner, A., Fried, I., Veshchev, I., Grossman, R. and Ram, Z,: A Non-Aggressive, Highly Efficient, Enzymatic Method for Dissociation of Human Brain-Tumors and Brain-Tissues to Viable Single-Cells., BMC Neurosci 17, 30, 2016
人	Human	淀粉樣蛋白原纖維 / Amyloid Fibrils	Collagenase Type 3: 0.02%； Deoxyribonuclease I: 0.001%	50mM Tris	Scherpelz, K., Lu, J.,Tycko, R. and Meredith, S.: Preparation of Amyloid Fibrils Seeded from Brain and Meninges, Methods Mol Biol 1345, 299, 2016
人		腦腫瘤 / Brain tumor	Collagenase Type 1: 0.04%； Hyaluronidase: 0.01%； Deoxyribonuclease I: 0.02%； Neutral Protease: 0.008%；	DMEM/F12	Hussein, D., Punjaruk, W., Storer, L., Shaw, L., Othman, R., Ottoman, R., Peet, A., Miller, S., Bandopadhyay, G., Heath, R., Kumari, R., Bowman, K., Braker, P., Rahman, R., Jones, G., Watson, S. and Lowe, J.: Pediatric Brain Tumor Cancer Stem Cells: Cell Cycle Dynamics, DNA Repair, and Etoposide Extrusion., Neuro Oncol 13, 70-83, 2011
人	Human	腫瘤 / Tumor	Collagenase Type 4: 0.1%； Hyaluronidase: 0.07%； Deoxyribonuclease I: 0.04%	see reference	Sauvageot, C., Weatherbee, J., Kesari, S., Winters, S., Barnes, J., Dellagatta, J., Ramakrishna, N., Stiles, C., Kung, A., Kieran, M. and Wen, P.: Efficacy of the HSP90 Inhibitor 17-AAG in Human Glioma Cell Lines and Tumorigenic Glioma Stem Cells.,Neuro Oncol Vol. 11,, , 109, 2009
人	Human	小膠質(zhì)細(xì)胞 / Microglia	Trypsin: 0.25%； Deoxyribonuclease I: .005%	DMEM/F12	Klegeris Andis, McGeer PatrickL: Chymotrypsin-like proteases contribute to human monocytic THP-1 cell as well as human microglial neurotoxicity, Glia 51, 56-64, 2005
人	Human	微血管 / Microvessels	Collagenase Type 4: 0.1%	DMEM	Gerhart, D. Z., Broderius, M. A., and Drewes, L. R.: Cutlured Human and Canine Endothelial Cells from Brain Microvessels, Brain Res Bull 21, 785, 1988
人	Human	神經(jīng)元 / Neuronal	Deoxyribonuclease I: 10 痢/ml	Tris-HCl, 50 mM, CaCl2, 2 mM	Roher, A.E., Palmer, K.C., Chau, V., and Ball, M.J.: Isolation and Chemical Characterization of Alzheimer's Disease Paired Helical Filament Cytoskeletons:Differentiation from Amyloid Plaque Core Protein, J Cell Biol 107, 2703, 1988
人	Human, 15-54 years	微血管 / Microvessels	Collagenase: 0.1%	Serum-free modified Lewis medium	Vinters, H.V., Reave, S., Costello, P., Girvin, J.P., and Moore, S.A.: Isolation and Culture of Cells Derived From Human Cerebral Microvessels, Cell Tissue Res 249, 657, 1987
昆蟲	Drosophilia	神經(jīng)元 / Neurons	Papain: 50 u/ml	Modified Dissecting Saline	Hadzic, T., Park, D., Abruzzi, K., Yang, L., Trigg, J., Rohs, R., Rosbash, M. and Taghert, P.: Genome-Wide Features of Neuroendocrine Regulation in Drosophila by the Basic Helix-Loop-Helix Transcription Factor DIMMED., Nucleic Acids Res 43, 2199-215, 2015
昆蟲	Drosophila	神經(jīng)元 / Neurons	Papain: 20 u/ml	Saline	Gu, H. and O'Dowd, D.: Whole Cell Recordings from Brain of Adult Drosophila., J Vis Exp 6, 248, 2007
昆蟲	Drosophila	神經(jīng)元 / Neurons	Papain: 4 u/ml	DMEM	Sicaeros, B. Campusano, J. and O'Dowd, D.: Primary Neuronal Cultures from the Brains of Late Stage Drosophila Pupae., J Vis Exp 4, 200, 2007
昆蟲	Drosophilia	肯揚(yáng) / Kenyon	Papain: 20 u/ml	Recording saline	Gu, H. and O'Dowd, D.: Cholinergic Synaptic Transmission in Adult Drosophila Kenyon Cells in Situ., J Neurosci 26, 265-72, 2006
猴	Monkey, rhesus	腦細(xì)胞 / Brain cells	Collagenase Type 2: 500 u/ml； Deoxyribonuclease I: 28 u/ml	HBSS	Marcondes MC, Burudi EM, Huitron-Resendiz S, Sanchez-Alavez M, Watry D, Zandonatti M, Henriksen SJ, Fox HS: Highly activated CD8(+) T cells in the brain correlate with early central nervous system dysfunction in simian immunodeficiency virus infection, J Immunol 167, 5429-38, 2001
小鼠	Adult and neonatal	小膠質(zhì)細(xì)胞，星形膠質(zhì)細(xì)胞，淋巴細(xì)胞 / Microglia, astrocytes, lymphocytes	Collagenase Type 2: 120 u/ml； Papain: 20 u/ml； Neutral Protease: 1.2 u/ml； Deoxyribonuclease I: 20 u/ml	EBSS	Calvo, Belen, Rubio, F., Fernandez, M. and Tranque, P.: Dissociation of Neonatal and Adult Mice Brain for Simultaneous Analysis of Microglia, Astrocytes and Infiltrating Lymphocytes by Flow Cytometry.,IBRO Rep Vol. 8,, , 36-47, 2020
小鼠	Mouse, adult	星形膠質(zhì)細(xì)胞，小膠質(zhì)細(xì)胞，內(nèi)皮細(xì)胞，少突膠質(zhì)細(xì)胞 / Astrocytes, microglia, endothelia, oligodendrocytes	Papain: 20 u/ml； Collagenase Type 1: 0.01%	HBSS	Swartzlander, D., Propson, N., Roy, E., Saito, T., Saido, T., Wang, B. and Zheng, H.: Concurrent Cell Type-Specific Isolation and Profiling of Mouse Brains in Inflammation and Alzheimer's Disease., JCI Insight 3, , 2018
小鼠	Mouse	神經(jīng)元 / Neurons	Papain: 10 u/ml； Deoxyribonuclease I: 10 u/ml	Artificial CSF	Kramer, D., Risso, D., Kosillo, ., Ngai, J. and Bateup, H.: Combinatorial Expression of Grp and Neurod6 Defines Dopamine Neuron Populations with Distinct Projection Patterns and Disease Vulnerability,eNeuro Vol. 5,, , 1, 2018
小鼠	Mouse, fetal and mature	星形膠質(zhì)細(xì)胞和神經(jīng)元 / Astrocytes and neurons	Papain: 7.5-20 u/ml	RPMI	Zhang, Y., Sloan, S., Clarke, L., Caneda, C., Plaza, C., Blumenthal, P., Vogel, H., Steinberg, G., Edwards, M., Li, G., Duncan, J., Cheshier, S., Shuer, L., Chang, E., Grant, G., Gephart, M. and Barres, B.: Purification and Characterization of Progenitor and Mature Human Astrocytes Reveals Transcriptional and Functional Differences with Mouse., Neuron 89, 37-53, 2016
小鼠	Mouse	脈絡(luò)叢 / Chloroid plexus	Collagenase Type 4: 400 u/ml	PBS	Baruch, K., Rosenzweig, N., Kertser, A., Deczkowska, A., Sharif, A., Spinrad, A., Tsitsou-Kampeli, A., Sarel, A., Cahalon, L. and Schwartz, M.: Breaking Immune Tolerance by Targeting Foxp3(+) Regulatory T Cells Mitigates Alzheimer's Disease Pathology., Nat Commun 6, 7967, 2015