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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(yǎng)和細(xì)胞生物學(xué)研究的原代細(xì)胞的分離和收集。盡管這些消化酶已經(jīng)普遍應(yīng)用多年，但是對其組織消化和細(xì)胞收集的作用機(jī)制卻沒有全面的理解。 由此導(dǎo)致的結(jié)果便是選擇一種消化方法大多是隨機(jī)選擇或基于過去的經(jīng)驗(yàn)，而不是在*理解消化原理的基礎(chǔ)上選擇的消化方案，以及如何根據(jù)具體實(shí)驗(yàn)?zāi)康男拚桨敢垣@得更好的實(shí)驗(yàn)結(jié)果。
細(xì)胞分離程序的目的是zui大程度的獲得功能活性的單個(gè)細(xì)胞，而影響結(jié)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(yǎng)液
6、所用的酶
7、消化酶中的雜質(zhì)
8、所用消化酶的工作濃度
9、溫度
10、孵育時(shí)間
通常前四個(gè)原因沒辦法選擇，而剩下的原因一般由經(jīng)驗(yàn)決定以獲取zui適實(shí)驗(yàn)結(jié)果。
通過查閱文獻(xiàn)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(shí)驗(yàn)數(shù)據(jù)搞得一頭霧水。這些巨大差異來源于組織胞外基質(zhì)的復(fù)雜而動態(tài)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(zhì)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(fā)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(jié)了目前常用組織消化酶的zui適消化和細(xì)胞收集知識和方法，描述了標(biāo)準(zhǔn)的實(shí)驗(yàn)步驟， 以便于制定一個(gè)*的細(xì)胞分離方案，同時(shí)列舉了一些相關(guān)的參考文獻(xiàn)。
下表列出多種組織的適宜消化酶（含參考文獻(xiàn)，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(jié)腸組織 | 內(nèi)皮組織 | 上皮組織 |
眼 | 心臟 | 腸 | 腎 |
肝 | 肺 | 淋巴結(jié) | 乳腺 |
其它組織 | 其他組織 | 肌組織 | 神經(jīng)組織 |
胰腺 | 腮腺 | 垂體 | 前列腺 |
生殖 | 動物鱗片 | 皮膚 | 脾臟 |
干細(xì)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(xiàn)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。
腎組織消化手冊
| 物種 | 詳細(xì)來源 | 細(xì)胞名稱 | 消化用酶 | 培養(yǎng)基 | 參考文獻(xiàn) |
| 鳥類 | 鳥類, 麻雀 | Tubule segments | Collagenase Type 2: 0.1% | HBSS | Goldstein D., Reddy V., and Plaga K.: Second Messenger Production in Avian Medullary Nephron Segments in Response to Peptide Hormones, Am J Physiol 276, R847, 1999 |
| 鳥類 | 鳥類, 雞 | Tubule and glomeruli | Collagenase Type 1: 0.1% | RPMI 1640 | Reich, C. and Bonar, R.: Separation of Avian Kidney Tubules and Glomeruli for In Vitro Culture, Meth Cell Sci 7, 97, 1982 |
| 牛 | 牛 | Papillary duct | Hyaluronidase: 0.2% | Keri's buffer | Husted, R., Hayashi, M., and Stokes, J.: Characteristics of Papillary Collecting Duct Cells in Primary Culture, Am J Physiol 255, F1160, 1988 |
| HEPES buffered saline | |||||
| 犬 | 犬 | Renal adipose derived cells | Collagenase Type 1: 0.3% | DMEM | Basu, J., Genheimer, C., Sangha, N., Quinlan, S., Guthrie, K., Kelley, R., Ilagan, R., Jain, D., Bertram, T. and Ludlow, J.: Organ Specific Regenerative Markers in Peri-Organ Adipose: Kidney., Lipids Health Dis 10, 171, 2011 |
| 犬 | 狗, 1-5 個(gè)月 | Proximal tubules | Collagenase Type 1: 0.15% | Krebs Ringer bicarbonate buffer | States, B., Reynolds, R., Lee, J., and Segal, S.: Cystine Uptake By Cultured Cells Originating From Dog Proximal Tubule Segments, In Vitro Cell Dev Biol 26, 105, 1990 |
| 犬 | 狗, 雜種狗, 20 公斤 | Proximal tubular | Deoxyribonuclease I: 0.0125% | (see reference) | Yau, C., Rao, L., and Silverman, M.: Sugar Uptake Into a Primary Culture of Dog Kidney Proximal Tubular Cells, Can J Physiol Pharmacol 63, 417, 1985 |
| 魚 | 美洲擬鰈 (鈍吻黃蓋鰈) | Renal tubule | Trypsin: 0.2% | CMF solution | Dickman, K. and Renfro, J.: Primary Culture of Flounder Renal Tubule Cells:Transepithelial Transport, Am J Physiol 251, 424, 1986 |
| 200 - 500 克 | (see reference) | ||||
| 豚鼠 | 豚鼠, 雄性, 100 克 | Single cells | Trypsin: 0.25% | CF salt solution | Phillips, H.: Dissociation of Single Cells from Lung or Kidney Tissue with Elastase, In Vitro 8, 101, 1972 |
| 倉鼠 | 倉鼠 | Papillary duct | Hyaluronidase: 0.2% | Keri's buffer | Husted, R., Hayashi, M., and Stokes, J.: Characteristics of Papillary Collecting Duct Cells in Primary Culture, Am J Physiol 255, F1160, 1988 |
| (包括SD大鼠, Wistar大鼠, 150-225 克, 兔, 牛) | HEPES buffered saline | ||||
| 人類 | 人類 | Renal adipose derived cells | Collagenase Type 1: 0.3% | DMEM | Basu, J., Genheimer, C., Sangha, N., Quinlan, S., Guthrie, K., Kelley, R., Ilagan, R., Jain, D., Bertram, T. and Ludlow, J.: Organ Specific Regenerative Markers in Peri-Organ Adipose: Kidney., Lipids Health Dis 10, 171, 2011 |
| 人類 | 人類, 成年人 | Renal tumor cells and proximal tubular epithelial | Collagenase Type 2: 0.1% | DMEM/F12 | Valente, M., Henrique, R., Costa, V., Jeronimo, C., Carvalho, F., Bastos, M., Guedes de Pinho, P. and Carvalho, M.: A Rapid and Simple Procedure for the Establishment of Human Normal and Cancer Renal Primary Cell Cultures from Surgical Specimens., PLoS ONE 6, e19337, 2011 |
| 人類 | 人類 | Renal cells | Collagenase Type 4: 300 u/ml | HBSS | Presnell, S., Bruce, A., Wallace, S., Choudhury, S., Genheimer, C., Cox, B., Guthrie, K., Werdin, E., Tatsumi-Ficht, P., Ilagan, R., Kelley, R., Rivera, E., Ludlow, J., Wagner, B., Jayo, M. and Bertram, T.: Isolation, Characterization, and Expansion Methods for Defined Primary Renal Cell Populations from Rodent, Canine, and Human Normal and Diseased Kidneys., Tiss Eng 17, 261, 2011 |
| Neutral Protease: see reference | |||||
| 人類 | 人類, 成年人 | Renal proximal tubule and cortical fibroblasts | Collagenase Type 2: 383 u/ml | DMEM/F-12 | Johnson, D., Saunders, H., Johnson, F., Huq, S., Field, M., and Pollock, C.: Cyclosporin exerts a direct fibrogenic effect on human tubulointerstitial cells: roles of insulin-like growth factor I, transforming growth factor beta1, and plaet-derived growth factor, J Pharmacol Exp Ther 289, 535-42, 1999 |
| 人類 | 人類 | Renal cortex | Trypsin: 0.1% | Tissue Culture Grade Water | McAteer, J, Kempson, S., and Evan, A: Culture of Human Renal Cortex Epithelial Cells, J Tiss Cul Meth 13, 143, 1991 |
| 人類 | 人類 | Papillary duct | Collagenase: 400 u/ml | Eagle's MEM-HEPES buffer w/L-glutamine | Trifillis, A. and Kahng, M.: Characterization of an In Vitro System of Human Renal Papillary Collecting Duct Cells, In Vitro Cell Dev Biol 26, 441, 1990 |
| 人類 | 人類 | Mesangial | Trypsin: 0.25% | DMEM/Ham's F-12 | Heieren, M., van der Woude, F., and Balfour Jr., H.: Cytomegalovirus Replicates Efficiently in Human Kidney Mesangial Cells, Proc Natl Acad Sci U S A 85, 1642, 1988 |
| 人類 | 人類, 成年人 | Tubular | Collagenase: 250 u/ml | PBS | Yang, A., Gould-Kostka, J., and Oberley, T.: In Vitro Growth and Differentiation of Human Kidney Tubular Cells on a Basement Membrane Substrate, In Vitro Cell Dev Biol 23 (1), 34, 1987 |
| 人類 | 人類, 成年人, 14-66 歲 | Tubular | Collagenase: 100 u/ml | Joklik's MEM | Trifillis, A., Regec, A., and Trump, B.: Isolation, Culture, and Charaterization of Human Renal Tubular Cells, J Urol 133, 324, 1985 |
| 人類 | 人類 | Malignant | Papain: 0.009% | Sacks solution | Hemstreet, G., Enoch, P., and Pretlow, T.: Tissue Disaggregation of Human Renal Cell Carcinoma with Further Isopyknic and Isokinetic Gradient Purification, Cancer Res 40, 1043, 1980 |
| Stromal | |||||
| 人類 | 人類, 24 初生和 夭折的 | Renal | Trypsin: 0.25% | See reference | De Oca, H., and Malinin, T.: Dispersion and C*tion of Renal Cells After Short-Term Storage of Kidneys, J Clin Microbiol 2, 243, 1975 |
| (包括兔) | |||||
| 猴 | 猴 | Kidney | Trypsin: 0.01% - 0.00001% | Eagle's MEM | Melnick, J., and Wallis, C.: Problems Related to the Use of Serum and Trypsin in the Growth of Monkey Kidney Cells, Dev Biol 37, 77, 1976 |
| 小鼠 | 小鼠 | Collecting duct epithelial | Collagenase Type 2: 0.1% | DMEM | DiRocco, D., Kobayashi, A., Taketo, M., McMahon, A. and Humphreys, B.: Wnt4/尾-catenin Signaling in Medullary Kidney Myofibroblasts., J Am Soc Nephrol 24, 1399, 2013 |
| 小鼠 | 小鼠 | Glomeruli | Collagenase Type 4: 0.1% | RPMI 1640 | Kabgani, N., Grigoleit, T., Schulte, K., Sechi, A., Sauer-Lehnen, S., Tag, C., Boor, P., Kuppe, C., Warsow, G., Schordan, S., Mostertz, J., Chilukoti, R., Homuth, G., Endlich, N., Tacke, F. and Weiskirchen, R.: Primary Cultures of Glomerular Parietal Epithelial Cells or Podocytes with Proven Origin., PLoS ONE 7, e34907, 2012 |
| 小鼠 | 小鼠, 10-14 week | Proximal tubule | Collagenase Type 1: 0.1% | See reference | Park, J., Pasupulati, R., Feldkamp, T., Roeser, N. and Weinberg, J.: Cyclophilin D and the Mitochondrial Permeability Transition in Kidney Proximal Tubules after Hypoxic and Ischemic Injury., Am J Physiol/Renal 301, F134, 2011 |
| 小鼠 | 小鼠, 胚胎 | Kidney | Collagenase Type 1: 0.25% | HBSS | Brown, A., Blank, U., Adams, D., Karolak, M., Fetting, J., Hill, B. and Oxburgh, L.:Isolation and Culture of Cells from the Nephrogenic Zone of the Embryonic Mouse Kidney., J Vis Exp 50, e2555, 2011 |
| Pancreatin: 1.0% | |||||
| Deoxyribonuclease I: 1 u/ml | |||||
| 小鼠 | 小鼠,成年的 | Kidney Sphere-Derived | Collagenase Type 4: 0.1% | Iscove's Modified Dulbecco's | Huang, Y., Johnston, P., Zhang, B., Zakari, A., Chowdhry, T., Smith, R., Marban, E., Rabb, H. and Womer, K.: Kidney-Derived Stromal Cells Modulate Dendritic and T Cell Responses., J Am Soc Nephrol 20, 831-41, 2009 |
| 小鼠 | 小鼠, 雄性, 8-16 周 | Renal tubular epithelial | Collagenase: 200 u/ml | HBSS | Breggia, A. and Himmelfarb, J.: Primary Mouse Renal Tubular Epithelial Cells have Variable Injury Tolerance to Ischemic and Chemical Mediators of Oxidative Stress., Oxid Med Cell Longev 1, 33, 2008 |
| Soybean Trypsin Inhibitor: see reference | |||||
| 小鼠 | 小鼠 | Proximal tubule | Collagenase Type 2: 0.013% | see reference | Wright, J., Morales, M., Sousa-Menzes, J., Ornellas, D., Sipes, J., Cui, Y., Cui, I., Hulamm, P., Cebotaru, V., Cebotaru, L., Guggino, W. and Guggino, S.: Transcriptional Adaptation to Clcn5 Knockout in Proximal Tubules of Mouse Kidney., Physiol Genomics 33, 341, 2008 |
| 小鼠 | 小鼠, 4 周 | Endothelial kidney | Collagenase Type 1: 0.1% | DMEM | Kondo, S., Scheef, E., Sheibani, N. and Sorenson, C.: PECAM-1 Isoform-Specific Regulation of Kidney Endothelial Cell Migration and Capillary Morphogenesis., Am J Physiol Cell Physiol 292, C2070, 2007 |
| 小鼠 | 小鼠, 成年的 | Cortex, proximal tubule | Collagenase: 0.15% | DMEM | Syal Ashu, Schiavi Susan, Chakravarty Sumana, Dwarakanath Vangipuram, Quigley Raymond, Baum Michel: Fibroblast growth factor-23 increases mouse PGE2 production in vivo and in vitro, Am J Physiol/Renal 290, F450-5, 2006 |
| 小鼠 | 小鼠 | Kidney Mesenchymal | Collagenase: 250 u/ml | DMEM/F12 | Plotkin, M. and Goligorsky, M.: Mesenchymal Cells from Adult Kidney Support Angiogenesis and Differentiate into Multiple Interstitial Cell Types including Erythropoietin-Producing Fibroblasts., Am J Physiol Renal Physiol 291, F902-12, 2006 |
| Neutral Protease: 33.3 u/ml | |||||
| 小鼠 | 小鼠, 10-12 周 | Kidney | Collagenase Type 4: 0.075% | DMEM | Dekel, B., Zangi, L., Shezen, E., Reich-Zeliger, S., Eventov-Friedman, S., Katchman, H., Jacob-Hirsch, J., Amariglio, N., Rechavi, G., Margalit, R. and Reisner, Y.: Isolation and Characterization of Nontubular sca-1+lin- Multipotent Stem/Progenitor Cells from Adult Mouse Kidney., J Am Soc Nephrol 17, 3300, 2006 |
| 小鼠 | 小鼠, 雄性, 12-16 周 | Renal proximal tubule cells | Collagenase Type 2: 0.1% | DMEM/F-12 | Cunningham, R., Xiaofei, E., Steplock, D., Shenolikar, S. and Weinman, E.: Defective PTH Regulation of Sodium-Dependent Phosphate Transport in NHERF-1-/- Renal Proximal Tubule Cells and Wild-Type Cells Adapted to Low-Phosphate Media., Am J Physiol Renal Physiol 289, F933, 2005 |
| Soybean Trypsin Inhibitor: 0.25% | |||||
| 小鼠 | 小鼠, 雄性, 2 個(gè)月 | Cortical collecting duct | Collagenase: 54-178 u/ml | MEM | Sindic, A., Velic, A., Basoglu, C., Hirsch, J., Edemir, B., Kuhn, M. and Schlatter, E.:Uroguanylin and Guanylin Regulate Transport of Mouse Cortical Collecting Duct Independent of Guanylate Cyclase C., Kidney Int 68, 1008, 2005 |
| Protease: see reference | |||||
| 小鼠 | 小鼠, 6 周 | Glomerular mesangial cells | Collagenase Type 4: 0.1% | RPMI 1640 | Radeke HH, Janssen-Graalfs I, Sowa EN, Chouchakova N, Skokowa J, Loscher F, Schmidt RE, Heeringa P, Gessner JE.: Opposite regulation of type II and III receptors for immunoglobulin G in mouse glomerular mesangial cells and in the induction of anti-glomerular basement membrane (GBM) nephritis, J Biol Chem 277(30), 27535, 2002 |
| 小鼠 | 小鼠, 3-7 個(gè)月 | Fibroblasts, mesangial, smooth muscle | Trypsin: 0.25% | DMEM | Bradshaw AD, Francki A, Motamed K, Howe C, Sage EH: Primary mesenchymal cells isolated from SPARC-null mice exhibit altered morphology and rates of proliferation, Mol Biol Cell 10, 1569-79, 1999 |
| Collagenase: see reference | |||||
| Soybean Trypsin Inhibitor: .05% | |||||
| 小鼠 | 小鼠 | Epithelial | Collagenase Type 4: 1% | DMEM /F-12 | Taub, M: Renal Tubule Cells,Cell & Tissue Culture: Laboratory Procedures Vol. 2,Doyle, A., Griffiths, J., and Newell, D., John Wiley & Sons, Ltd., 14B:6.1, 1995 |
| 小鼠 | 小鼠,單純的 SJL (H-2S) | Proximal tubular epithelial | Deoxyribonuclease I: 15 礸/ml | RPMI 1640 | Haverty, T., Kelly, C., Hines, W., Amenta, P., Watanabe, M., Harper, R., Kefalides, N., and Neilson, E.: Characterization of a Renal Tubular Epithelial Cell Line Which Secretes the Autologous Target Antigen of Autoimmune Experimental Interstitial Nephritis, J Cell Biol 107, 1359, 1988 |
| 豬 | 豬, 雜種 | Kidney | Collagenase Type 1: 0.1% | HBSS | Heussner, A. and Dietrich, D.: Primary Porcine Proximal Tubular Cells as an Alternative to Human Primary Renal Cells in Vitro: an Initial Characterization., BMC Cell Biol 14, 55, 2013 |
| 兔 | 兔 | Renal proximal tubular | Collagenase Type 1: 60 u/ml | DMEM/F12 | Nowak, G. and Bakajsova, D.: Assessment of Mitochondrial Functions and Cell Viability in Renal Cells Overexpressing Protein Kinase C Isozymes., J Vis Exp 71, e4301, 2013 |
| 兔 | 兔, 新西蘭 | Proximal tubule cells | Collagenase Type 4: 0.005% | DMEM/F-12 | Taub, M.: Primary Kidney Proximal Tubule Cells., Methods Mol Biol 290, 231, 2005 |
| Soybean Trypsin Inhibitor: 0.00025% | |||||
| 兔 | 兔, 新西蘭 白, 雌性 | Nephron segments | Collagenase Type 2: 0.05% | Eagle's MEM | Schafer, J., Watkins, M., Li, L., Herter, P., Haxelmans, S., and Schlatter, E.: A Simplified Method for Isolation of Large Numbers of Defined Nephron Segments, Am J Physiol 273, F650, 1997 |
| Deoxyribonuclease I: 50 u/ml | |||||
| 兔 | 兔, 新西蘭 白, 雄性, 1.5 - 2.0 公斤 | Duct | Soybean Trypsin Inhibitor: 0.025% | Hank's Solution with calcium and HEPES | Naray-Fejes-Toth, A., and Fejes-Toth, G.: Immunoelection and Culture of Cortical Collecting Duct Cells, J Tiss Cul Meth 13, 179, 1991 |
| 兔 | 兔, 新西蘭 掰,雌性, 2-3 公斤 | Renal proximal tubules | Deoxyribonuclease I: 70 u/ml | Modified DME-F12 | Rodeheaver, D., Aleo, M., and Schnellmann, R.: Differences in Enzymatic and Mechanical Isolated Rabbit Renal Proximal Tubules: Comparison in Long-Term Incubation, In Vitro Cell Dev Biol 26, 898, 1990 |
| 兔 | 兔 | Papillary duct | Hyaluronidase: 0.2% | Keri's buffer | Husted, R., Hayashi, M., and Stokes, J.: Characteristics of Papillary Collecting Duct Cells in Primary Culture, Am J Physiol 255, F1160, 1988 |
| (包括, SD 大鼠和Wistar大鼠, 150-225克, 馬, 牛) | HEPES buffered saline | ||||
| 兔 | 兔, 新西蘭, 白色, 2-3 公斤 | MTALH cells | Collagenase: 0.1% | DMEM | Allen, M., Nakao, A., Sonnenburg, W., Burnatowska-Hledin, M., Speilman, W., and Smith, W.: Immunodissection of Cortical and Medullary Thick Ascending Limb Cells From Rabbit Kidney, Am J Physiol 255, F704, 1988 |
| RCCT cells | |||||
| 兔 | 兔, 新西蘭, 白色 | Collecting tubule | Trypsin: 0.05% | Kreb's Ringer buffer | Grenier, F., and Smith, W.: Formation of 6-keto-PGF-1aby Collecting Tubule Cells Isolated from Rabbit Renal Papillae, Prostaglandins 16, 759, 1978 |
| 兔 | 兔, 新西蘭, 白色, 雌性, 2.5 公斤 | Papillary collecting duct | Collagenase: 0.2% | (see reference) | Dworzack, D., and Grantham, J.: Preparation of Renal Papillary Collecting Duct Cells for Study, Nat Neurosci 8, 191, 1975 |
| 大鼠 | SD大鼠, 胎兒 | Fetal kidney | Collagenase Type 4: 0.1% | MEM | Gupta, A., Jadhav, S., Tripathy, N. and Nityanand, S.: Fetal Kidney Cells Can Ameliorate Ischemic Acute Renal Failure in Rats through Their Anti-Inflammatory, Anti-Apoptotic and Anti-Oxidative Effects., PLoS ONE 10, e0131057, 2015 |
| 大鼠 | Lewis大鼠, 雄性 | Renal adipose derived cells | Collagenase Type 1: 0.3% | DMEM | Basu, J., Genheimer, C., Sangha, N., Quinlan, S., Guthrie, K., Kelley, R., Ilagan, R., Jain, D., Bertram, T. and Ludlow, J.: Organ Specific Regenerative Markers in Peri-Organ Adipose: Kidney., Lipids Health Dis 10, 171, 2011 |
| 大鼠 | 大鼠 | Proximal tubule | Collagenase Type 2: 0.1% | see reference | Panico, C., Luo, Z., Damiano, S., Artigiano, F., Gill, P. and Welch, W.: Renal Proximal Tubular Reabsorption is Reduced in Adult Spontaneously Hypertensive Rats: Roles of Superoxide and Na+/H+ Exchanger 3., Hypertension 54, 1291, 2009 |
| 大鼠 | SD大鼠, 雄性, 200-250克 | Medullary thick ascending limb | Collagenase: 0.1% | HEPES-saline | Silva, G., Beierwaltes, W., and Garvin, J.: Extracellular ATP stimulates NO production in rat thick ascending limb, Hypertension 47, 563-7, 2006 |
| 大鼠 | Wistar大鼠, 300克 | Renal proximal tubules | Collagenase Type 2: 0.2% | DMEM/F12 | Deng Aihua, Miracle Cynthia M, Lortie Mark, Satriano Joseph, Gabbai Francis B, Munger Karen A, Thomson Scott C, Blantz Roland C: Kidney oxygen consumption, carbonic anhydrase, and proton secretion, Am J Physiol/Renal 290, F1009-15, 2006 |
| 大鼠 | 大鼠, Wistar, 雄性, 300-350 克 | Renal proximal tubules | Collagenase Type 2: 0.2% | DMEM/F-12 | Deng, A., Miracle, C., Suarez, J., Lortie, M., Satriano, J., Thomson, S., Munger, K. and Blantz, R.: Oxygen Consumption in the Kidney: Effects of Nitric Oxide Synthase Isoforms and Angiotensin II., Kidney Int 68, 723, 2005 |
| 大鼠 | 大鼠 | Cortical tubule cells | Collagenase: 0.6% | DMEM | Arystarkhova, E., Wetzel R., and Sweadner K.: Distribution and Oligomeric Association of Splice Forms of Na(+)-K(+)-ATPase Regulatory Gamma-subunit in Rat Kidney, Am J Physiol/Renal 282(3), F393, 2002 |
| 大鼠 | SD大鼠, 雄性 275-300 克 | Glomerular mesangial cells | Collagenase Type 1: 250 u/ml | DMEM | Amiri Farhad, Shaw Sean, Wang Xiaodan, Tang Jie, Waller Jennifer L, Eaton Douglas C, Marrero Mario B: Angiotensin II activation of the JAK/STAT pathway in mesangial cells is altered by high glucose, Kidney Int Suppl 61, 1605-16, 2002 |
| 大鼠 | SD大鼠, 1 周 | Cortical collection duct | Collagenase Type 2: 0.15% | DMEM/Ham's F12 | Valencia L., Bidet M., Martial S., Sanchez E., Melendez E., Tauc M., Poujeol C., Martin D., Namorado M., Reyes J., and Poujeol P.: Nifedipine-activated Ca(2+) Permeability in Newborn Rat Cortical Collecting Duct Cells in Primary Culture, Am J Physiol Cell Physiol280(5), C1193, 2001 |
| 大鼠 | SD大鼠, 250-300克 | Microvessels | Collagenase Type 2: 0.1% | PSS | Li, N., Teggatz, E., Li, P., Allaire, R., and Zou, A.: Formation and Actions of Cyclic ADP-Ribose in Renal Microvessels, Microvasc Res 60, 149, 2000 |
| 大鼠 | SD大鼠, 250-300 克 | Renal | Collagenase Type 2: 0.2% | see reference | Mattson D., and Wu F.: Nitric Oxide Synthase Activity and Isoforms in Rat Renal Vasculature, Hypertension 35, 337, 2000 |
| 大鼠 | SD大鼠, 雄性, 5-6 周 | Renal tubules | Collagenase: 0.1% | see reference | Miyata, N., Park, F., Li, X., and Cowley, A.: Distribution of Angiotensin AT1 and AT2 Receptor Subtypes in the Rat Kidney, Am J Physiol 277, F437, 1999 |
| 大鼠 | SD大鼠, 225-250 g | Glomerular mesangial cells | Collagenase Type 1: 250 u/ml | DMEM | Amiri F, Garcia R: Regulation of angiotensin II receptors and PKC isoforms by glucose in rat mesangial cells, Am J Physiol 276, F691-9, 1999 |
| 大鼠 | SD大鼠, 雄性 | Proximal tubule suspensions | Collagenase Type 4: 0.1% | HEPES buffer | Eitle E, Hiranyachattada S, Wang H, Harris PJ: Inhibition of proximal tubular fluid absorption by nitric oxide and atrial natriuretic peptide in rat kidney, Am J Physiol 274, C1075-80, 1998 |
| Pronase E: 2 u/ml | |||||
| 大鼠 | 大鼠, SD, 雄性 | Nephron segments | Collagenase Type 2: 0.05% | Eagle's MEM | Schafer, J., Watkins, M., Li, L., Herter, P., Haxelmans, S., and Schlatter, E.: A Simplified Method for Isolation of Large Numbers of Defined Nephron Segments, Am J Physiol 273, F650, 1997 |
| Deoxyribonuclease I: 50 u/ml | |||||
| 大鼠 | 大鼠, 12-周大 | Renal | Collagenase: 0.1% | DMEM | Ishikawa, S., Kusaka, I., Higashiyama, M., Nagasaka, S., Saito, T., Honda, K., and Saito, T.: Cellular Signaling and Proliferative Action of AVP in Mesangium of SHR: Effect of Low Density Lipoprotein, Nat Neurosci 50, 1506, 1996 |
| 大鼠 | 大鼠, 雄性, 150-200 克 | Epithelial | Protease: 0.1% | HBSS/CMF | Elliget, K., and Trump, B.: Primary Cultures of Normal Rat Kidney Proximal Tubule Epithelial Cells for Studies of Renal Cell Injury, In Vitro Cell Dev Biol 27, 739, 1991 |
| Proximal tubule | |||||
| 大鼠 | 大鼠, 成年的 | Inner medullary collecting duct | Collagenase Type 2: 0.1% | PBS | Brion, L., Schwartz, J., Lachman, H., Zavilowitz, B., and Schwartz, G.: Development of H+ Secretion by Cultured Renal Inner Medullary Collecting Duct Cells, Am J Physiol 257, F486, 1989 |
| Papillae | |||||
| 大鼠 | Wistar大鼠, 雌性, 100-150 克 | Glomerular mesangial | Collagenase: 0.025% | RPMI 1640 | Wang, J., Kester, M., and Dunn, M.: The Effects of Endotoxin on Plaet-Activating Factor Synthesis in Cultured Rat Glomerular Mesangila Cells, Biochim Biophys Acta 969, 217, 1988 |
| 大鼠 | Wistar大鼠, 雄性, 150 - 200 克 | Renal target | Collagenase: 1.0% (also 0.7%) | Eagle's MEM | Barlet-Bas, C., Khadouri, C., Marsey, S., and Doucet, A.: Sodium-Independent In VitroInduction of Na+,K+-ATPase by Aldosterone in Renal Target Cells: Permissive Effect of Triiodothyronine, Proc Natl Acad Sci U S A 85, 1707, 1988 |
| 大鼠 | Wistar大鼠, 雄性, 成年的 | Proximal tubules | Collagenase: 0.2% | Krebs-Henseleit buffer | Gesek, F., Wolff, D., and Strandhov, J.: Improved Separation Method for Rat Proximal and Distal Renal Tubules, Am J Physiol 253, F358, 1987 |
| 大鼠 | Wistar大鼠, 300-400克 | Proximal tubules | Collagenase: 0.15 % | Krebs Henseleit solution | Vinay, P., Gougoux, A., and Lemieux, G.: Isolation of a Pure Suspension of Rat Proximal Tubules, Am J Physiol 241, F403, 1981 |
| 大鼠 | 大鼠 | Fibroblasts | Trypsin: up to 0.25% | Dulbecco-Vogt MEM | Wallach, D., Anderson, W., and Pastan, I.: Activation of Adenylate Cyclase in Cultured Fibroblasts by Trypsin, J Biol Chem 253, 24, 1978 |
| Kidney |
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