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Worthington組織消化指南
引言
組織消化酶主要用于組織培養和細胞生物學研究的原代細胞的分離和收集。盡管這些消化酶已經普遍應用多年，但是對其組織消化和細胞收集的作用機制卻沒有全面的理解。 由此導致的結果便是選擇一種消化方法大多是隨機選擇或基于過去的經驗，而不是在*理解消化原理的基礎上選擇的消化方案，以及如何根據具體實驗目的修正消化方案以獲得更好的實驗結果。
細胞分離程序的目的是zui大程度的獲得功能活性的單個細胞，而影響結果的原因有很多，包括（不限于）以下：
1、組織類型
2、物種來源
3、動物年齡
4、遺傳改造情況（如knockouts）
5、所用的消化培養液
6、所用的酶
7、消化酶中的雜質
8、所用消化酶的工作濃度
9、溫度
10、孵育時間
通常前四個原因沒辦法選擇，而剩下的原因一般由經驗決定以獲取zui適實驗結果。
通過查閱文獻來選擇*組織消化酶和zui適消化條件的研究人員通常被互相矛盾的實驗數據搞得一頭霧水。這些巨大差異來源于組織胞外基質的復雜而動態的特性，以及過去常用的粗制、未提純的組織分離酶。另外，組織胞外基質由各種蛋白、糖蛋白、脂類和糖脂組成，這些成分的組成比例在不同物種、不同組織、甚至不同發育（年齡）之間存在明顯的差異，這些因素都極大的影響了消化方案的制定。目前通常使用的粗制酶，如鏈霉蛋白酶、NF1:250、以及含有幾種其他不同濃度蛋白酶的膠原酶、以及各種多糖酶、核酶和脂酶。
本指南總結了目前常用組織消化酶的zui適消化和細胞收集知識和方法，描述了標準的實驗步驟， 以便于制定一個*的細胞分離方案，同時列舉了一些相關的參考文獻。
下表列出多種組織的適宜消化酶（含參考文獻，以組織類型和物種分類，詳見鏈接）
脂肪組織 | 腎上腺組織 | 骨組織 | 腦組織 |
軟骨組織 | 結腸組織 | 內皮組織 | 上皮組織 |
眼 | 心臟 | 腸 | 腎 |
肝 | 肺 | 淋巴結 | 乳腺 |
其它組織 | 其他組織 | 肌組織 | 神經組織 |
胰腺 | 腮腺 | 垂體 | 前列腺 |
生殖 | 動物鱗片 | 皮膚 | 脾臟 |
干細胞 | 胸腺 | 甲狀腺/甲狀旁腺 | 扁桃體 |
腫瘤 |
|
|
|
注：使用Worthington公司消化酶的文獻不限于上表鏈接中所列，一般而言，Worthington公司的組織消化酶可用于大多組織類型的消化。
腦組織分離手冊
| 物種 | 詳細來源 | 細胞名稱 | 消化用酶 | 培養基 | 參考文獻 |
| 牛 | 牛 | 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 | 牛 | 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 | 牛 | 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 | 牛, 成年的 | Cerebral artery | 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 |
| Endothelial | |||||
| 牛 | 牛 | 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 | 牛 | 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 | 牛 | Endothelial | Collagenase Type 2: 0.2% | Dulbecco's PBS | Goetz, I.., Warren, J., Estrada, C., Roberts, E., and Krause, D.: Long-Term C*tion of Arterial and Capillary Endothelium From 成年的 Bovine Brain, In Vitro Cell Dev Biol 21, 172, 1985 |
| Brain arteries | |||||
| 牛 | Calf | Oligodendroglia | 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 |
| (also lamb) | Neural | ||||
| 豚鼠 | 豚鼠, 200-400 g | Neurons | Trypsin: 0.06-0.08% | PIPES saline | Kay, A.R., and Wong, R.K.S.: Isolation of Neurons Suitable for Patch-Clamping from 成年的 Mammalian Central Nervous Systems, J Neurosci Methods 16, 227, 1986 |
| 人類 | 人類 | Tumor | Collagenase Type 4: 0.1% | 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 |
| Hyaluronidase: 0.07% | |||||
| Deoxyribonuclease I: 0.04% | |||||
| 人類 | 人類 | Microglia | Trypsin: 0.25% | 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 |
| Deoxyribonuclease I: .005% | |||||
| 人類 | 人類 | 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 |
| 人類 | 人類 | Neuronal | Deoxyribonuclease I: 10 礸/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 |
| 人類 | 人類, 15-54 歲 | Microvessels | Collagenase: 0.1% | Serum-free modified Lewis medium | Vinters, H.V., Reave, S., Coslo, 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 | 果蠅 | Neurons | Papain: 20 u/ml | Saline | Gu, H. and O'Dowd, D.: Whole Cell Recordings from Brain of 成年的 Drosophila., J Vis Exp 6, 248, 2007 |
| 昆蟲 | 果蠅 | 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 |
| 猴 | 猴, 獼猴 | Brain cells | Collagenase Type 2: 500 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 |
| Deoxyribonuclease I: 28 u/ml | |||||
| 小鼠 | 小鼠 | 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 |
| 小鼠 | 小鼠 | Brain-sequestered leukocytes | Collagenase Type 4: 0.05% | RPMI | Ryg-Cornejo, V., Ioannidis, L. and Hansen, D.: Isolation and Analysis of Brain-Sequestered Leukocytes from Plasmodium Berghei ANKA-Infected Mice., J Vis Exp71, e50112, 2013 |
| Deoxyribonuclease I: 2 u/ml | |||||
| 小鼠 | 小鼠 | Cerebral pericytes | Collagenase Type 2: 0.1% | DMEM | Shah. G., Price, T., Banks, W., Morofuji, Y., Kovac, A., Ercal, N., Sorenson, C., Shin, E., Sheibani, N.: Pharmacological Inhibition of Mitochondrial Carbonic Anhydrases Protects Mouse Cerebral Pericytes from High Glucose-Induced Oxidative Stress and Apoptosis., J Pharmacol Exp Ther 344, 637-45, 2013 |
| Deoxyribonuclease I: 30 u/ml | |||||
| 小鼠 | 小鼠, P13-15 | Glial | Papain: 20 u/ml | EBSS | Haseleu, J., Anlauf, E., Blaess, S., Endl, E. and Derouiche, A.: Studying Subcellular Detail in Fixed Astrocytes: Dissociation of Morphologically Intact Glial Cells (DIMIGs)., Front Cell Neurosci 7, 54, 2013 |
| Deoxyribonuclease I: 0.0005% | |||||
| 小鼠 | 小鼠, 成年的 | Neurons | PDS kit: with modifications | DMEM/F12 | Saxena, A., Wagatsuma, A., Noro, Y., Kuji, T., Asaka-Oba, A., Watahiki, A., Gurnot, C., Fagiolini, M., Hensch, T. and Carninci, P.: Trehalose-Enhanced Isolation of Neuronal Sub-Types from 成年的 Mouse Brain., Biotechniques 52, 381, 2012 |
| 小鼠 | 小鼠, 1-2 天 | Oligodendrocytes, dorsal root ganglia | Papain: 0.15% | DMEM | O'Meara, R., Ryan, S., Colognato, H. and Kothary, R.: Derivation of Enriched Oligodendrocyte Cultures and Oligodendrocyte/Neuron Myelinating Co-Cultures from Post-Natal Murine Tissues., J Vis Exp 54, 3324, 2011 |
| Deoxyribonuclease I: 0.006% | |||||
| 小鼠 | 小鼠, 6-12周 | Vascular smooth muscle cells | Papain: 0.05% | PBS | Chung, W., Farley, J., Swenson, A., Barnard, J., Hamilton, G., Chiposi, R. and Drummond, H.: Extracellular Acidosis Activates ASIC-like Channels in Freshly Isolated Cerebral Artery Smooth Muscle Cells., Am J Physiol Cell Physiol 298, C1198, 2010 |
| Collagenase Type 4: 0.15% | |||||
| Elastase: 0.05% | |||||
| 小鼠 | 小鼠, 4-6 天 | Neurons | PDS kit: with modifications | HBSS | Lee, H., Greene, L., Mason, C. and Manzini, M.: Isolation and Culture of Post-Natal Mouse Cerebellar Granule Neuron Progenitor Cells and Neurons., J Vis Exp23, 990, 2009 |
| 小鼠 | 小鼠, 產后的 | Astrocytes | Trypsin: 0.25% | HBSS | Sher, F., Rossler, R., Brouwer, N., Balasubramaniyan, V., Boddeke, E. and Copray, S.: Differentiation of Neural Stem Cells Into Oligodendrocytes: Involvement of the Polycomb Group Protein Ezh2., Stem Cells 26, 2875, 2008 |
| Deoxyribonuclease I: 1,000 u/ml | |||||
| 小鼠 | 小鼠, 初生的 | Neurons | Papain: 20 u/ml | Neurobasal | Fasano, C., Thibault, D. and Trudeau, L.: Culture of Postnatal Mesencephalic Dopamine Neurons on an Astrocyte Monolayer,Current Protocols in Neuroscience Vol. 44,, , 3.21.1, 2008 |
| 小鼠 | 小鼠, 胚胎和出生后 | Cortical neurons | Papain: 4-10 u/ml | Neurobasal | Hilgenberg, L. and Smith, M.: Preparation of Dissociated Mouse Cortical Neuron Cultures., J Vis Exp 10, 562, 2007 |
| 小鼠 | 小鼠 | Neurosperes | PDS kit: per instructions | DMEM/F12 | Klein C, Butt SJ, Machold RP, Johnson JE, and Fishell G.: Cerebellum- and forebrain-derived stem cells possess intrinsic regional character, Development132, 4497, 2005 |
| 小鼠 | 小鼠, 1 天 | Neural progenitor | PDS kit: per instructions | see reference | Seaberg, R., Smukler, S. and Van der Kooy, D.: Intrinsic Differences Distinguish Transiently Neurogenic Progenitors from Neural Stem Cells in the Early Postnatal Brain., Dev Biol 278, 71, 2005 |
| 小鼠 | 小鼠 | Granule cell precursors, pre-neoplastic and tumor cells | Papain: 10 u/ml | Neurobasal/B27 | Oliver, T., Read, T., Kessler, J., Mehmeti, A., Wells, J., Huynh, T., Lin, S. and Wechsler-Reya, R.: Loss of Patched and Disruption of Granule Cell Development in a Pre-Neoplastic Stage of Medulloblastoma., Development 132, 2425, 2005 |
| Deoxyribonuclease I: 250 u/ml | |||||
| 小鼠 | 小鼠 | Cortical neurons | PDS kit: per instructions | Neurobasal | Hernandez, F., Perez, M., Lucas, J., Mata, A., Bhat, R. and Avila, J.: Glycogen Synthase Kinase-3 plays a Crucial Role in Tau Exon 10 Splicing and Intranuclear Distribution of SC35. Implications for Alzheimer's Disease., J Biol Chem 279, 3801, 2004 |
| 小鼠 | 小鼠, 3 天 | Microglia | Papain: 90 u/ml | Eagle's MEM | Nishioku T, Hashimoto K, Yamashita K, Liou SY, Kagamiishi Y, Maegawa H, Katsube N, Peters C, von Figura K, Saftig P, Katunuma N, Yamamoto K, Nakanishi H: Involvement of cathepsin E in exogenous antigen processing in primary cultured murine microglia, J Biol Chem 277, 4816, 2002 |
| Deoxyribonuclease I: 2000 u/ml | |||||
| 小鼠 | 小鼠, SD, 8-12 周 | Microglia | Trypsin: 0.125% | RPMI-1640 | O'Donnell SL, Frederick TJ, Krady JK, Vannucci SJ, Wood TL: IGF-I and microglia/macrophage proliferation in the ischemic mouse brain, Glia 39, 85, 2002 |
| Collagenase Type 2: 0.01% | |||||
| Deoxyribonuclease I: .005% | |||||
| 小鼠 | 小鼠, 2-5天 | Postnal substantia nigra | PDS kit: per instructions | see reference | Smeyne Michelle, Smeyne RichardJ: Method for culturing postnatal substantia nigra as an in vitro model of experimental Parkinson's disease, Brain Res Brain Res Protoc 9, 105-11, 2002 |
| 小鼠 | 小鼠, 胚胎 | Cortical progenitors | PDS kit: per instructions | Serum free medium | Estivill-Torrus, G., Pearson, H., Van Heyningen, V., Price, D. and Rashbass, P.:Pax6 is Required to Regulate the Cell Cycle and the Rate of Progression from Symmetrical to Asymmetrical Division in Mammalian Cortical Progenitors.,Development 129, 455, 2002 |
| 小鼠 | 小鼠, 1 天 | Neurons and glia | PDS kit: per instructions | Neurobasal | Martin-Aparicio, E., Yamamoto, A., Hernandez, F., Hen, R., Avila, J. and Lucas, J.: Proteasomal-Dependent Aggregate Reversal and Absence of Cell Death in a Conditional Mouse Model of Huntington's Disease., J Neurosci 21, 8772, 2001 |
| 小鼠 | 小鼠 | Cerebellar granule cell precursors | Papain: 0.435% | EBSS | Miyazawa K, Himi T, Garcia V, Yamagishi H, Sato S, and Ishizaki Y.: A role for p27/Kip1 in the control of cerebellar granule cell precursor proliferation, J Neurosci 20, 5756, 2000 |
| Deoxyribonuclease I: 0.05% | |||||
| 小鼠 | 小鼠 | Astrocytes | Trypsin: 0.25% | DMEM | Lim, D, and Alvarez-Buylla, A.: Interaction Between Astrocytes and 成年的 Subventricular Zone Precursors Stimulates Neurogenesis, Proc Natl Acad Sci U S A 96, 7526, 1999 |
| 小鼠 | 小鼠, 初生的 | Hippocampal cells | Papain: 10 u/ml | DMKM | Jun K, Choi G, Yang SG, Choi KY, Kim H, Chan GC, Storm DR, Albert C, Mayr GW, Lee CJ, Shin HS: Enhanced hippocampal CA1 LTP but normal spatial learning in inositol 1,4,5-trisphosphate 3-kinase(A)-deficient mice, Learn Mem 5, 317-30, 1998 |
| 小鼠 | 小鼠, SWR or CF1, 1-3 個月 | Papillae, taste receptor | Pronase E: 0.15% | Carbonate-Phosphate buffer | Spielman, A., Mody, I., Brand, J., Whitney, G., MacDonald, J., and Salter, M.: A Method for Isolating and Patch-Clamping Single Mammalian Taste Receptor Cells,Brain Res 503, 326, 1989 |
| (see reference) | |||||
| 小鼠 | 小鼠, 0-30 天 | Neural | Trypsin NF 1:250: 50 0.25% | BSS | Shrier, B., Wilson, S., and Nirenberg, M.: Cultured Cell Systems and Methods for Neurobiology, Vol. 32,, , 765, 1974 |
| 綿羊 | 羔羊 | Oligodendroglia | 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 |
| (還有 牛犢) | Neural | ||||
| 豬 | 小型豬, 尤卡坦半島 (尤卡坦半島野豬), 4-6 個月 | Microvascular | Collagenase: 0.1% | HBSS | Robinson, D.H., Kang,Y., Deschner, S.H., and Nielsen, T.B.: Morphologic Plasticity and Periodicity : Porcine Cerebral Microvascular Cells in Culture, In Vitro Cell Dev Biol 26, 169, 1990 |
| 大鼠 | 大鼠, SD, 19-21 天 | Suprachiasmatic nucleus neurons | Papain: 100 u/ml | MEM | Cao, R., Li, A., Cho, H., Lee, B. and Obrietan, K.: Mammalian Target of Rapamycin Signaling Modulates Photic Entrainment of the Suprachiasmatic Circadian Clock., J Neurosci 30, 6302, 2010 |
| 大鼠 | 大鼠, 食魚動物, 7-21 個月 | Hippocampal neurons | Papain: 0.2% | Hibernate A | Chen, N., Newcomb, J., Garbuzova-Davis, S., Davis Sanberg, C., Sanberg, P. and Willing, A.: Human Umbilical Cord Blood Cells Have Trophic Effects on Young and Aging Hippocampal Neurons in Vitro., Aging Dis 1, 173, 2010 |
| 大鼠 | 大鼠, SD, 7 天 | Cerebellar granule neurons | PDS kit: per instructions | PBS | Tanaka, S., Shaikh, I., Chiocca, E. and Saeki, Y.: The Gs-Linked Receptor GPR3 Inhibits the Proliferation of Cerebellar Granule Cells During Postnatal Development., PLoS ONE 4, e5922, 2009 |
| 大鼠 | 大鼠, neonatal | Astrocytes | Papain: 20 u/ml | EBSS | Shigetomi, E. and Khakh, B.: Measuring Near Plasma Membrane and Global Intracellular Calcium Dynamics in Astrocytes., J Vis Exp 26, 1142, 2009 |
| 大鼠 | 大鼠,初生的 | Hippocampal neurons | Papain: 10 u/ml | EBSS | Richler Esther, Chaumont Severine, Shigetomi Eiji, Sagasti Alvaro, Khakh Baljit S: Tracking transmitter-gated P2X cation channel activation in vitro and in vivo,Nat Methods 5, 87-93, 2008 |
| 大鼠 | 大鼠, SD | Hippocampal | Trypsin: 0.1% | Neurobasal | Akanda, N., Tofighi, R., Brask, J., Tamm, C., Elinder, F. and Ceccali, S.:Voltage-Dependent Anion Channels (VDAC) in the Plasma Membrane Play a Critical Role in Apoptosis in Differentiated Hippocampal Neurons but not in Neural Stem Cells., Cell Cycle 7, 3225, 2008 |
| 大鼠 | 大鼠, neonatal | Dura mater | Collagenase Type 1: 0.075% | DMEM | Peptan, I., Hong, L. and Evans, C.: Multiple Differentiation Potentials of Neonatal Dura Mater-Derived Cells., Neurosurgery 60, 346, 2007 |
| 大鼠 | 大鼠, E18 | Hippocampal neurons | Papain: 0.2% | Hibernate | Jekabsons MB, Nicholls DG: Bioenergetic analysis of cerebellar granule neurons undergoing apoptosis by potassium/serum deprivation, Cell Death Differ 13, 1595-610, 2006 |
| 大鼠 | 大鼠, 威斯塔, 1-3 day | Hippocampal neurons | Trypsin: 0.05% | DMEM | Velasco Myrian, Garci璦 Esperanza, Onetti Carlos G: Glucose deprivation activates diversity of potassium channels in cultured rat hippocampal neurons,Cell Mol Neurobiol 26, 307-19, 2006 |
| 大鼠 | 大鼠, E19 | Hippocampal neuron | Papain: 10 u/ml | MEM | Khakh Baljit S, Fisher James A, Nashmi Raad, Bowser David N, Lester Henry A:An angstrom scale interaction between plasma membrane ATP-gated P2X2 and alpha4beta2 nicotinic channels measured with fluorescence resonance energy transfer and total internal reflection fluorescence microscopy, J Neurosci 25, 6911-20, 2005 |
| 大鼠 | 大鼠, 威斯塔, 14天 | Visual cortical | PDS kit: per instructions | EBSS | Mizoguchi Y, Kanematsu T, Hirata M, Nabekura J: A rapid increase in the total number of cell surface functional GABAA receptors induced by brain-derived neurotrophic factor in rat visual cortex, J Biol Chem 278, 44097, 2003 |
| 大鼠 | 大鼠, 威斯塔, 雄性 | Cerebral endothelial | Collagenase Type 3: 0.2% | MEM | Floris S, Van den Born J, van der Pol SM, Dijkstra CD, De Vries HE: Heparan sulfate proteoglycans modulate monocyte migration across cerebral endothelium, J Neuropathol Exp Neurol 62, 780, 2003 |
| 大鼠 | 大鼠, SD, E18 | Cortical cells | Papain: 20 u/ml | EBSS | Behar, T, Smith, S, Kennedy, R, Mckenzie, J, Maric, I and Barker, J: GABA(B) receptors mediate motility signals for migrating embryonic cortical cells, Cereb Cortex 11, 744-53, 2001 |
| Deoxyribonuclease I: .005% | |||||
| 大鼠 | 大鼠, 威斯塔, 250-300 g | Cerebral artery smooth muscle cells | Papain: 1.5 mg/ml | Physiological Salt Solution | Brzezinska AK, Gebremedhin D, Chilian WM, Kalyanaraman B, Elliott SJ.:Peroxynitrite reversibly inhibits Ca(2+)-activated K(+) channels in rat cerebral artery smooth muscle cells, Am J Physiol Heart Circ Physiol 278, H1883, 2000 |
| Collagenase Type 4: 1.5 mg/ml | |||||
| 大鼠 | 大鼠, SD, E16 | Cortical neurons | PDS kit: with modifications | Neurobasal Medium | Varney MA, Cosford ND, Jachec C, Rao SP, Sacaan A, Lin FF, Bleicher L, Santori EM, Flor PJ, Allgeier H, Gasparini F, Kuhn R, Hess SD, Velicelebi G, Johnson EC:SIB-1757 and SIB-1893: selective, noncompetitive antagonists of metabotropic glutamate receptor type 5, J Pharmacol Exp Ther 290, 170, 1999 |
| 大鼠 | 大鼠, 威斯塔, 初生的 | Hippocampal cells | Papain: 1 mg/ml | DMEM | Boehm S, Betz H: Somatostatin inhibits excitatory transmission at rat hippocampal synapses via presynaptic receptors, J Neurosci 17, 4066, 1997 |
| 大鼠 | 大鼠, SD, 食魚動物 | Hippocampal neurons | Papain: 0.2% | HibernateA/B27 | Brewer, G.J.: Isolation and culture of 成年的 rat hippocampal neurons of any age, J Neurosci Methods 71, 143, 1997 |
| 大鼠 | 大鼠, E18 | Hippocampal and cortical neurons | PDS kit: per instructions | Neurobasal media | Naeve GS, Ramakrishnan M, Kramer R, Hevroni D, Citri Y, Theill LE: Neuritin: a gene induced by neural activity and neurotrophins that promotes neuritogenesis,Proc Natl Acad Sci U S A 94, 2648-53, 1997 |
| 大鼠 | 大鼠, SD, 初生 | Glial | Trypsin: 0.0625% | MEM, sterile | Pixley, S.K.: The Olfactory Nerve Contains Two Populations of Glia, In Vitro 5, 269, 1992 |
| 大鼠 | 大鼠, 初生 | Astrocytes | Trypsin: 0.25% | DMEM, HBSS | Holzwarth, J., Glaum, S., and Miller, R.: Activation of Endothelin Receptors by Sarafotoxin Regulates Ca2+ Homeostasis in Cerebellar Astrocytes, Glia 5, 239, 1992 |
| 大鼠 | 大鼠, 60-72 小時大的 | Fibroblasts | Trypsin: 0.2% | HEPES buffered DMEM | Acheson, A., Barker, P., Alderson, R., Miller, F., and Murphy, R.: Detection of Brain-Derived Neurotrophic Factor-like Activity in Fibroblasts and Schwann Cells: Inhibition by Antibodies to NGF, Neuron 7, 265, 1991 |
| 大鼠 | 大鼠, SD, 成年的 | Endothelial | Collagenase Type 2: 0.5% | Medium 199 | Doron, D., Jacobowitz, D., Heldman, E., Feurerstein, G., Pollard, H., and Hallenbeck, J.: Extracellular Matrix Permits the Expression of Von Willebrand's Factor, Uptake of Di-I-Acetylated Low Density Lipoprotein and Secretion of Prostacyclin in Cultures of Endothelial Cells from Rat Brain Microvessels, In Vitro Cell Dev Biol 27A, 689, 1991 |
| 大鼠 | 大鼠, SD, 5天 | Hippocampal neurons | Trypsin: 0.2% | HBSS | Mattson, M., and Kater, S.: Development and Selective Neurodegeneration in Cell Cultures From Different Hippocampal Regions, Brain Res 490, 110, 1989 |
| Deoxyribonuclease I: 10 ug/ml | |||||
| 大鼠 | 大鼠, SD, 雌性 | Fetal rat brain | Collagenase Type 4: 0.1% | DMEM | Matsuda, M.: Serum Proteins Enhance Aggregate Formation of Dissociated Fetal Rat Brain Cells in an Aggregating Culture, In Vitro Cell Dev Biol 24 (10), 1031, 1988 |
| 大鼠 | 大鼠, 白化病, 成年的 和初生的 | Cerebral cortices | Trypsin: 0.25% | PBS | Giulian, D. and Baker, T.J.: Characterization of Ameboid Microglia Isolated From Developing Mammalian Brain, J Neurosci 6, 2163, 1986 |
| 大鼠 | 大鼠, 威斯塔, 初生 | Germinal matrix | Trypsin: 0.25% | HBSS | Goldman, J.E., Geier, S.S., and Hirano, M.: Differentiation of Astrocytes and Oligodendrocytes From Germinal Matrix Cells in Primary Culture, J Neurosci 6, 52, 1986 |
| 大鼠 | 大鼠,胎兒, 18-20天 | Hippocampal neurons | Trypsin: 0.25% | HBSS, CMF | Bartlett, W. and Banker, G.: An Electron Microscopic Study of the Development of Axons and Dendrites by Hippocampal Neurons in Culture. I. Cells Which Develop Without Intercellular Contacts, J Neurosci 4, 1944, 1984 |
| 大鼠 | 大鼠, 胎兒, 18 天 | Hippocampi | Trypsin: 0.1% | HBSS | Rothman, S.: Synaptic Release of Excitatory Amino Acid Neurotransmitter Mediates Anoxic Neuronal Death, J Neurosci 4 (7), 1884, 1984 |
| 大鼠 | 大鼠, SD, 懷孕19-20天的 | Neural | Trypsin: 0.25% | DMEM | Ahmed, Z., Walker, P., and Fellows, R.: Properties of Neurons from Dissociated Fetal Rat Brain in Serum-Free Culture, J Neurosci 3 (12), 2448, 1983 |
| 大鼠 | 大鼠, SD, 胎兒 | Cerebral cortex | Deoxyribonuclease I: 0.001% | HEPES | Peterfreund, R. and Vale, W.: High Molecular Weight Somatostatin Secretion by Cultured Rat Brain Cell, Brain Res 239, 463, 1982 |
| Hypothalmus | |||||
| 大鼠 | Wistar京都大鼠, 雄性, 100 - 200 克 | Endothelial | Collagenase Type 2: 0.05% | HBSS | Diglio, C.A., Grammas, P., Filiberto Giacomelli, M.S., and Wiener, J.: Primary Culture of Rat Cerebral Microvascular Endothelial Cells, Lab Invest 46, 554, 1982 |
| Cerebral | |||||
| 大鼠 | 大鼠, 胎兒 | Cortical | Trypsin: 0.25% | Puck's D1 (see reference) | Swaiman, K., Neale, E., Fitzgerald, S., and Nelson, P.: A Method for Large-scale Production of Mouse Brain Cortical Cultures, Brain Res 255, 361, 1982 |
| 大鼠 | 大鼠, 威斯塔,懷孕的 | Glial | Trypsin: 0.05% | Eagle's MEM/DMEM | Abney, E., Bartlett, P., and Raff, M.: Astrocytes, Ependymal Cells, and Oligodendrocytes Develop on Schedule in Dissociated Cell Cultures of Embryonic Rat Brain, Dev Biol 83, 301, 1981 |
| 大鼠 | 大鼠, SD, 一個月大的 | Capillary endothelium | Neutral Protease: 0.1% | Medium 199 | Bowman, P., Betz, A., Ar, D., Wolinsky, J., Penney, J., Shivers, R., and Goldstein, G.: Primary Culture of Capillary Endothelium From Rat Brain, In Vitro 17 (4), 353, 1981 |
| Pericytes | |||||
| 大鼠 | 大鼠, 威斯塔, 雄性, 300-500 克 | Microvessels | Collagenase Type 2: 0.75% | Ringers-HEPES buffer | Williams, S., Gillis, J., Matthews, M., Wagner, R., and Bitensky, M.: Isolation and Characterization of Brain Endothelial Cells: Morphology and Enzyme Activity, J Neurochem 35 (2), 374, 1980 |
| Endothelial | |||||
| 大鼠 | 大鼠, 威斯塔, 成年的, 170克 | Endothelial | Trypsin: 0.5% | BSS | Phillips, P., Kumar, P., Kumar, S., and Waghe, M.: Isolation And Characterization of Endothelial Cells From Rat And Cow Brain White Matter, J Anat 129, 261, 1979 |
| 大鼠 | 大鼠, 豪斯曼, 18 天 | Hippocampal neurons | Trypsin: 0.1% | HBSS | Banker, G., and Cowan, M.: Rat Hippocampal Neurons in Dispersed Cell Culture,Brain Res 126, 397, 1977 |
| 大鼠 | 大鼠, SD, 2 天 | Pineal | Trypsin: 0.25% | DMEM, MEM | Rowe, V., Neale, E., Avins, L., Guroff, G., and Schrier, B.: Pineal gland cells in culture. Morphology, Biochemistry, Differentiation, and co-culture with sympathetic neurons, Exp Cell Res 104, 345, 1977 |
| 貝類動物 | 旋節螺trivolvis | Neurons | Trypsin: 0.2% | Saline, Sterile | Hadley, R.D., Bodnar, D.A., and Kater, S.B.: Formation of Electrical Synapses Between Isolated, Cultured Helisoma Neurons Requires Mutual Neurite Elongation,J Neurosci 5, 3145, 1985 |
| Buccal ganglia |
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