胰腺組織消化手冊

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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，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)具體實驗?zāi)康男拚桨敢垣@得更好的實驗結(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、孵育時間

通常前四個原因沒辦法選擇，而剩下的原因一般由經(jīng)驗決定以獲取zui適實驗結(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驗數(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)的實驗步驟，以便于制定一個*的細(xì)胞分離方案，同時列舉了一些相關(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。

胰腺組織消化手冊

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)
牛	牛	Duct epithelial	Collagenase: 0.1%	HEPES	Cotton, C., and Al-Nakkash, L.: Isolation and Culture of Bovine Pancreatic Duct Epithelial cells, Am J Physiol 272, G1328, 1997
牛	牛 (歐洲牛)	Ductal	Neutral Protease: 0.05%	EBSS	Sato, T., Sato, M., Hudson, E., and Jones, R.: Characterization of Bovine Pancreatic Ductal Cells Isolated by a Perfusion-Digestion Technique, In Vitro 19, 651, 1983
牛	牛	Plaets	Trypsin:	(see reference)	Stiles, G., and Lefkowitz, R.: Hormone-Sensitive Adneylate Cyclase, J Biol Chem 257 (11), 6287, 1982
犬	犬	Buccal ganglia; SLT muscle	Trypsin: 0.2%	DMEM	Zoran, M., Doyle, R. and Haydon, P.: Target Contact Regulates the Calcium Responsiveness of the Secretory Machinery During Synaptogenesis, Neuron 6, 145, 1991
犬	犬, 成年雜種, 15-25 公斤	Islets	Collagenase Type 4: 600-1100 u/ml	RPMI 1640	Noel, J., Rabinovitch, A., Olson, L., Kyriakides, G., Miller, J., and Mintz, D.: A Method for Large-Scale, High-Yield Isolation of Canine Pancreatic Islets of Langerhans, Metabolism 31 (2), 184, 1982
犬	犬, 成年雜種, 15-25 公斤	Islets	Deoxyribonuclease I: 10 ug/ml	RPMI 1640
魚	魚,絲鱸, 3個月大 0.5 克	Islets	Collagenase: 0.12 - 0.46 u/ml	RPMI 1640	Schrezenmeir, J., Laue, C., Sternheim,E., Wolbert, K., Darquy, S., Chicheportiche, D., Kirchgessner, J., and Reach, G.: Long-Term Function of Single-Cell Preparations of Piscine Principal Islets in Hollow Fibers, Transplant Proc 24 (6), 2941, 1992
豚鼠	豚鼠	Acinar	Collagenase Type 3: 60 u/ml	Kreb's Ringer	Schultz, G., Sarras, Jr, M., Gunther, G., Hull, B., Alicea, H., Gorelick, F., and Jamieson, J.: Guinea Pig Pancreatic Acini Prepared with Purified collagenase, Exp Cell Res 130, 49, 1980
豚鼠	Hartley豚鼠, 白化變種, 雄性, 350-400 克	Acinar	Soybean Trypsin Inhibitor: 0.01%	Kreb's Ringer	Gardner, J., Conlon, T., Klaeveman, H., Adams, T., and Ondetti, M.: Action of Cholecystokinin and Cholinergic Agents on Calcium Transport in Isolated Pancreatic Acinar Cells, J Clin Invest 56, 366, 1975
豚鼠	豚鼠	Exocrine	Hyaluronidase: 0.15% - 0.2%	Kreb's Ringer	Amsterdam, J., and Jamieson, J.: Structural and Functional Characterization of Isolated Pancreatic Exocrine Cells, Proc Natl Acad Sci U S A 69 (10), 3028, 1972
馬	馬	Islets	Collagenase Type 4: 1.3% - 2.0%	HBSS	Feldman, J., and Chapman, B.: Preparation of Islets of Langerhans from Rabbits and Hamsters by the Collagenase Digestion Technique, Acta Diabetol 12, 208, 1975
人類	人類	Acinar	CLSPA: 200 u/ml	See Reference	Cane, M., Sutton, R. and Criddle, D.: Isolation of Human Pancreatic Acinar Cells From Rat and Human Pancreas, Pancreapedia 20, 1, 2011
人類	人類	Acinar	Soybean Trypsin Inhibitor: 0.01%	See Reference
人類	人類	Pancreatic cancer stem cells	Collagenase Type 4: 200 u/ml	medium 199	Li, C., Heidt, D., Dalerba, P., Burant, C., Zhang, L., Adsay, V., Wicha, M., Clarke, M. and Simeone, D.: Identification of Pancreatic Cancer Stem Cells., Cancer Res 67, 1030, 2007
人類	人類, 25-55 歲大	Islets	Collagenase: 0.4%	HBSS	Contractor, H., Johnson, P., Chadwick, D., Robertson, G., and London, N.: The Effect of UW Solution and Its Components on the Collagenase Digestion of Human and Porcine Pancreas, Cell Transplant 4 (6), 615, 1995
人類	(包括豬)	Islets	Collagenase: 0.4%	HBSS
人類	人類	Islets	Collagenase: 0.2%	Eurocollins solution	Watt, P., Mullen, Y., Benhamou, P., Hober, C., Nomura, Y., Watanabe, Y., Passaro, E., Zinner, M., and Brunicardi, F.: Simplified Semiautomated Method For Isolating Islets From the Human Pancreas, Transplant Proc 26 (2), 582, 1994
人類	人類	Islets	Deoxyribonuclease I: 200 u/ml	Eurocollins solution
人類	人類, 成人器官捐獻(xiàn)者	Islets	Collagenase: 0.6%	Eurocollins solution	Warnock, G., Rajotte, R., Evans, M., Ellis, D., DeGroot, T., and Dawidson, I.: Isolation of Islets of Langerhans Following Cold Storage of Human Pancreas, Transplant Proc XIX (4), 3466, 1987
人類	人類	Islets	Collagenase Type 4: 0.8%	HBSS	Izumi, R., Konishi, K., Ueno, K., Shimizu, K., Hirosawa, H., Takahashi, N., and Miyazaki, I.: Isolation of Human Pancreatic Islets from Cryopreserved Pancreas,Transplant Proc XVII, 383, 1985
人類	人類	Islets	Collagenase (1 or 4): 0.60%	HBSS	Gray, D., McShane, P., Grant, A., and Morris, P.: A Method for Isolation of Islets of Langerhans from the Human Pancreas, Diabetes 33, 1055, 1984
人類	人類, 嬰兒, a克e 1 天-1 歲	Islets	Collagenase: 170-210 u/ml	HBSS	Sutherland, D., Matas, A., Steffes, M., and Najarian, J.: Infant Human Pancreas: A Potential Source of Islet Tissue for Transplantation, Diabetes 25 (12), 1123, 1976
猴	猴, 3-5 公斤	Islets	Hyaluronidase: 0.05%	HBSS	Scharp, D., Murphy, J., Newton, W., Ballinger, W., and Lacy, P.: Application of an Improved Isolation Technique for Islet Transplantation in Primates and Rats,Transplant Proc 7, 739, 1975
小鼠	小鼠	Islets	Collagenase: 0.1-0.25%	HBSS	Koh, D., Moody, M. and Jo, J.: Collection of Islets of Langerhans using an Equilibrium Method., Biotechniques 55, 34-7, 2013
小鼠	小鼠, 8-12 周	Pancreas organoid	Collagenase: 0.012%	DMEM	Huch, M., Bonfanti, P., Boj, S., Sato, T., Loomans, C., Van de Wetering, M., Sojoodi, M., Li, V., Schuijers, J., Gracanin, A., Ringnalda, F., Begthel, H., Hamer, K., Mulder, J., Van Es, J. and De Koning, E.: Unlimited In Vitro Expansion of Adult Bi-Potent Pancreas Progenitors Through the Lgr5/R-Spondin Axis., EMBO J 32, 2708-21, 2013
小鼠	小鼠, 8-12 周	Pancreas organoid	Neutral Protease: 0.012%	DMEM
小鼠	小鼠, 胚胎	Pancreatic progenitor	Neutral Protease: 0.125%	DMEM	Greggio, C., De Franceschi, F., Figueiredo-Larsen, M., Gobaa, S., Ranga, A., Semb, H., Lutolf, M. and Grapin-Botton, A.: Artificial Three-Dimensional Niches Deconstruct Pancreas Development In Vitro., Development 140, 4452-62, 2013
小鼠	小鼠	Islets	Collagenase: 0.2%	RPMI 1640	Bertera, S., Balamurugan, A., Bottino, R., He, J. and Trucco, M.: Increased Yield and Improved Transplantation Outcome of Mouse Islets with Bovine Serum Albumin., J Transplant 2012, 856386, 2012
小鼠	小鼠, 10 周	Islets	Collagenase: 0.2%	RPMI 1640	Kobayashi, T., Yamaguchi, T., Hamanaka, S., Kato-Itoh, M., Yamazaki, Y., Ibata, M., Sato, H., Lee, Y., Usui, J., Knisely, A., Hirabayashi, M. and Nakauchi, H.: Generation of Rat Pancreas in Mouse by Interspecific Blastocyst Injection of Pluripotent Stem Cells., Cell142, 787, 2010
小鼠	小鼠, 6-8 個月	Islets	Collagenase Type 4: 0.1%	HBSS	Taguchi, Y., Tasaki, Y., Terakado, K., Kobayashi, K., Machida, T. and Kobayashi, T.:Impaired Insulin Secretion from the Pancreatic Islets of Hypothyroidal Growth-Retarded Mice., J Endocrinol 206, 195-204, 2010
小鼠	小鼠, 成年的, 雄性	Ancinar	CLSPA: see reference	DMEM	Ji, B., Gaiser, S., Chen, X., Ernst, S. and Logsdon, C.: Intracellular Trypsin Induces Pancreatic Acinar Cell Death but not NF-KappaB Activation., J Biol Chem 284, 17488, 2009
小鼠	小鼠, 成年的, 雄性	Ancinar	Soybean Trypsin Inhibitor: 0.001%	DMEM
小鼠	小鼠, 3-4 周, 20-24 克	Islets	Collagenase Type 4: 0.2%	RPMI 1540	Huang, H., Xie, Q., Kang, M., Zhang, B., Zhang, H., Chen, J., Zhai, C., Yang, D., Jiang, B. and Wu, Y.: Labeling Transplanted Mice Islet with Polyvinylpyrrolidone Coated Superparamagnetic Iron Oxide Nanoparticles for In Vivo Detection By Magnetic Resonance Imaging., Nanotechnology 20, 365101, 2009
小鼠	小鼠	Islets	Collagenase: 1,000 u/ml	RPMI 1640	Li, D., Yuan, Y., Tu, H., Liang, Q. and Dai, L.: A Protocol for Islet Isolation from Mouse Pancreas., Nat Protoc 4, 1649, 2009
小鼠	小鼠	Islets	Collagenase: 0.14%	RPMI 1640	Carter, J., Dula, S., Corbin, K., Wu, R. and Nunemaker, C.: A Practical Guide to Rodent Islet Isolation and Assessment., Biol Proced Online 11, 3, 2009
小鼠	小鼠	Islets	Collagenase: 0.03-0.08%	RPMI 1640	Szot, G., Koudria, P. and Bluestone, J.: Murine Pancreatic Islet Isolation., J Vis Exp 7, 255, 2007
小鼠	小鼠, 6-10 個月	Pancreatic ductal	CLSPA: 50 u/ml	DMEM	Wang, Y., Soyombo, A., Shcheynikov, N., Zeng, W., Dorwart, M., Marino, C., Thomas, P. and Muallem, S.: Slc26a6 Regulates CFTR Activity In Vivo to Determine Pancreatic Duct HCO3- Secretion: Relevance to Cystic Fibrosis., EMBO J 25, 5049, 2006
			Hyaluronidase: 400 u/ml
			Soybean Trypsin Inhibitor: 0.02%
小鼠	小鼠, 成年的	Acinar	CLSPA: 200 u/ml	see reference	Voronina, S., Barrow, S., Gerasimenko, O., Petersen, O. and Tepikin, A.: Effects of Secretagogues and Bile Acids on Mitochondrial Membrane Potential of Pancreatic Acinar Cells: Comparison of Different Modes of Evaluating Delta Psi., J Biol Chem 279, 27327, 2004
小鼠	小鼠, 11-12 周	Islets	Collagenase Type 4: 0.2%	RPMI	Astrof, S., Crowley, D., George, E., Fukuda, T., Sekiguchi, K., Hanahan, D. and Hynes, R.: Direct Test of Potential Roles of EIIIA and EIIIB Alternatively Spliced Segments of Fibronectin in Physiological and Tumor Angiogenesis., Mol Cell Biol 24, 8662-70, 2004
小鼠	小鼠	Pancreatic islet	Collagenase Type 4: 0.2%	HBSS	Haefliger Jacques-Antoine, Tawadros Thomas, Meylan Laure, Gurun SabineLe, Roehrich Marc-Estienne, Martin David, Thorens Bernard, Waeber Gerard: The scaffold protein IB1/JIP-1 is a critical mediator of cytokine-induced apoptosis in pancreatic beta cells, J Cell Sci 116, 1463-9, 2003
小鼠	小鼠	Islets	Collagenase Type 4: 0.2%	HBSS	Wu Yulian, Han Bing, Luo Hongyu, Roduit Raphael, Salcedo TheodoraW, Moore PaulA, Zhang Jun, Wu Jiangping: DcR3/TR6 effectively prevents islet primary nonfunction after transplantation, Diabetes 52, 2279-86, 2003
小鼠	小鼠, BALB/c, 6-8 周-old, either sex	Acinar	Collagenase: 0.1%	Waymouth's MB	Kurup, S., and Bhonde, R.: Analysis and Optimization of Nutritional Set-up for Murine Pancreatic Acinar Cells, JOP 3 (1), 8, 2002
小鼠	小鼠	Islets	Collagenase Type 2: 0.2%	CF Medium	Koster, J., Marshall, B., Ensor, N., Corbett, J., and Nichols, C.: Targeted Overativity of Cell K_ATP Channels Induces Profound Neonatal Diabetes, Cell 100, 645, 2000
小鼠	小鼠	Pancreatic islets	Collagenase Type 4: 0.4%	Gey's BSS	Strowski M, Parmar R, Blake A, Schaeffer J: Somatostatin inhibits insulin and glucagon secretion via two receptors subtypes: an in vitro study of pancreatic islets from somatostatin receptor 2 knockout mice, Endocrinology 141, 111-7, 2000
小鼠	小鼠	Acinar cells and acini	Collagenase Type 1: see reference	see reference	Toivola, D., Ku, N., Ghori, N., Lowe, A., Michie, S. and Omary, M.: Effects of Keratin Filament Disruption on Exocrine Pancreas-Stimulated Secretion and Susceptibility to Injury., Exp Cell Res 255, 156, 2000
小鼠	小鼠	Acinar cells and acini	CLSPA: see reference	see reference
小鼠	小鼠, 白話病變, 25 克	Acinar	CLSPA: see reference	see reference	Fogarty, K., Kidd, J., Tuft, R. and Thorn, P.: A Bimodal Pattern of InsP(3)-Evoked Elementary Ca(2+) Signals in Pancreatic Acinar Cells., Biophys J 78, 2298, 2000
小鼠	小鼠	Duct	Papain: 25 u/ml	DMEM /F-12	Githens, S: Pancreatic Duct Epithelial Cells,Cell & Tissue Culture: Laboratory Procedures Vol. 1,Doyle, A., Griffiths, J., and Newell, D., John Wiley and Sons, Ltd., 12B:12.1, 1995
小鼠	小鼠	Acinar	Collagenase: 100 u/ml	HEPES	Jauch, P., Peterson, O., and Lauger, P.: Electrogenic Properties of the Na-Alanine Cotransporter in Acinar Cells, J Membr Biol 94, 99, 1986
小鼠	小鼠 (C57BL/6J-ob/ob), 9-12 周, 雄性	Islets	Hyaluronidase: 0.5%	Kreb's Ringer bicarbonate buffer	Dalpe-Scott, M., Heick, H., and Begin-Heick, N.: Secretion in the Obese (ob/ob) Mouse. The Effect of Oxytetracycline on Insulin Release, Diabetes 32, 932, 1983
小鼠	小鼠, 雄性, 18-24 克	Acinar	CLSPA: 70-90 u/ml	Krebs-Henseleit	Burnham DB, Williams JA: Effects of carbachol, cholecystokinin, and insulin on protein phosphorylation in isolated pancreatic acini, J Biol Chem 257, 10523-8, 1982
小鼠	小鼠, 雄性, 18-24 克	Acinar	Soybean Trypsin Inhibitor: 0.01%	Krebs-Henseleit
小鼠	小鼠, 7-10 周	Islets	Collagenase: see reference	DMEM	Yesil, P, Michel, M., Chwalek, K., Pedack, S., Jany, C., Ludwig, B., Bornstein, S. and Lammert, E.: A New Collagenase Blend Increases the Number of Islets Isolated from Mouse Pancreas., Islets 1, 185,
豬	豬, 少年的	Islets	Collagenase: see reference	Univ of Wisconsin solution	Van der Burg Michael P M, Graham John M: Iodixanol Density Gradient Preparation in University of Wisconsin solution for porcine islet purification, ScientificWorldJournal 3, 1154-9, 2003
豬	豬, 3 個月, 15-20 公斤	Acinar	Collagenase Type 3: 200 u/ml	RPMI 1640	Zhao, X., Han, J., and Tang, C.: Primary Culture of Porcine Pancreatic Acinar Cells, JOP2 (2), 78, 2001
豬	豬, 1-3 歲, 1.5-2.0 公斤, 任一性別	Islets	Collagenase: 0.25%	HBSS	Korbutt, G., Elliott, J., Ao, Z., Smith, D., Warnock, G., and Rajotte, R.: Large Scale Isolation, Growth, and Function of Porcine Neonatal Islet Cells, J Clin Invest 97 (9), 2119, 1996
豬	豬, 雌性	Islets	Collagenase: 0.1%	HBSS	Brandhorst, D., Brandhorst, H., Hering, B., Federlin, K., and Bretzel, R.: Islet Isolation from the Pancreas of Large Mammals and Humans: 10 歲s of Experience, Exp Clin Endocrinol 103, 3, 1995
豬	豬	Islets	Collagenase: 0.1%-0.2%	HBSS	Johnson, P., van Suylichem, P., Roberts, D., Vos-Scheperkeuter, G., White, S., van Schlifgaarde, R., London, N.: Design of a Simple, in vitro Method for Evaluation of the Efficiency of Crude Clostridium histolyticum Collagenase and its Components for Porcine Islet Isolation, Xenotransplantation 2, 165, 1995
豬	豬, <2 歲, 200-250 公斤	Islets	Collagenase: 0.1%	HBSS	Heiser, A., Ulrichs, K., and Muller-Ruchholtz, W.: Isolation of Porcine Pancreatic Islets: Low Trypsin Activity During the Isolation Procedure Guarantees Reproducible High Islet Yields, J Clin Lab Anal 8, 407, 1994
豬	豬, 10-36 個月s, 200-300 公斤	Islets	Collagenase: 0.2%	HBSS	Ricordi, C., Socci, C., Davalli, A., Staudacher, C., Baro, P., Vertova, A., Sassi, I., Gavazzi, F., Pozza, G., and Di Carlo, V.: Isolation of the Elusive Pig Islet, Surgery 107 (6), 688, 1990
豬	豬	Acinar	Collagenase: 100 u/ml	Saline	Iwatsuki, N., and Peterson, O.: Action of Tetraethylammonium on Calcium-Activated Potassim Channels in Pig Pancreatic Acinar Cells Studied by Patch-Clamp Single-Channel and Whole-Cell Current Recording, J Membr Biol 86, 139, 1985
兔	兔, 新西蘭白, 雄性/Fe雄性, 2-3 公斤	Acinar	Hyaluronidase: 0.2%	Kreb's Ringer bicarbonate buffer	Renckens, B., Schrijen, J., Swarts, H., DePont, J., and Bonting, S.: Role of Calcium in Exocrine Pancreatic Secretion. IV. Calcium Movements in Isolated Acinar Cells of Rabbit Pancreas, Biochim Biophys Acta 544, 338, 1978
兔	兔, 包括馬	Islets	Collagenase Type 4: 1.3% - 2.0%	HBSS	Feldman, J., and Chapman, B.: Preparation of Islets of Langerhans from Rabbits and Hamsters by the Collagenase Digestion Technique, Acta Diabetol 12, 208, 1975
大鼠	Wistar大鼠, 300克	Islets	Collagenase Type 4: 0.2%	CMRL 1066	Verga Falzacappa, C., Mangialardo, C., Raffa, S., Mancuso, A., Piergrossi, P., Moriggi, G., Piro, S., Stigliano, A., Torrisi, M., Brunetti, E., Toscano, V. and Misiti, S.: The Thyroid Hormone T3 Improves Function and Survival of Rat Pancreatic Islets During In Vitro Culture., Islets 2, 96, 2010
大鼠	大鼠	Acinar	CLSPA: 100 u/ml	DMEM	Williams, J.: Isolation of Rodent Pancreatic Acinar Cells and Acini by Collagenase Digestion, Pancreapedia , , 2010
大鼠	大鼠, 成年的	Islets	Collagenase Type 1: 450 u/ml	Leibowitz L-15	Williams, J., Huang, H., Kover, K., Moore, W., Berkland, C., Singh, M., Smirnova, I., MacGregor, R. and Stehno-Bit, L.: Reduction of Diffusion Barriers in Isolated Rat Islets Improves Survival, but not Insulin Secretion or Transplantation Outcome., Organogenesis6, 115, 2010
大鼠	大鼠, 雄性, 7-11 周	Islets	Collagenase Type 4: see reference	RPMI 1640	Getty-Kaushik, L., Richard, A., Deeney, J., Shirihai, O. and Corkey, B.: The CB1 Antagonist Rimonabant Decreases Insulin Hypersecretion in Rat Pancreatic Islets, Obesity17, 1856, 2009
大鼠	大鼠	Islets	Collagenase Type 1: 450 u/ml	RPMI 1640	MacGregor, R., Williams, S., Tong, P., Kover, K., Moore, W. and Stehno-Bit, L.:Small Rat Islets are Superior to Large Islets in In Vitro Function and in Transplantation Outcomes., Am J Physiol/Endo 290, E771, 2006
大鼠	Wistar大鼠, 雄性, 250-400克	Pancreatic islets	Collagenase: 0.75%	RPMI 1640	Tian XH, Xue WJ, Ding XM, Pang XL, Teng Y, Tian PX, and Feng XS: Small intestinal submucosa improves islet survival and function during in vitro culture, World J Gastroenterol 11, 7378, 2005
大鼠	大鼠, 250-350 克	Pancreatic islet	Collagenase Type 4: 0.2%	HBSS	Haefliger Jacques-Antoine, Tawadros Thomas, Meylan Laure, Gurun SabineLe, Roehrich Marc-Estienne, Martin David, Thorens Bernard, Waeber Gerard: The scaffold protein IB1/JIP-1 is a critical mediator of cytokine-induced apoptosis in pancreatic beta cells, J Cell Sci 116, 1463-9, 2003
大鼠	大鼠	Pancreatic acini	CLSPA: 30 u/ml	M199	Blinman TA, Gukovsky I, Mouria M, Zaninovic V, Livingston E, Pandol SJ, Gukovskaya AS: Activation of pancreatic acinar cells on isolation from tissue: cytokine upregulation via p38 MAP kinase, Am J Physiol Cell Physiol 279, C1993-2003, 2000
			Collagenase Type 4: 30 u/ml
			Soybean Trypsin Inhibitor: 0.01%
大鼠	SD大鼠, 雄性	Ancinar	CLSPA: see reference	DMEM	Ji B, Kopin AS, Logsdon CD: Species differences between rat and mouse CCKA receptors determine the divergent acinar cell response to the cholecystokinin analog JMV-180, J Biol Chem 275, 19115-20, 2000
大鼠	SD大鼠, 雄性	Ancinar	Soybean Trypsin Inhibitor: 0.01%	DMEM
大鼠	大鼠	Islets	Collagenase: 126 - 196 u/ml	HBSS	Verspohl, E., and Wienecke, A.: The Role of Protein Kinase C in the Desensitization of Rat Pancreatic Islets to Chrolinergic stimulation, J Endocrinol 159, 287, 1998
大鼠	大鼠	Islets	Collagenase: 0.5 - 0.9%	HBSS	Takaki, R and Ono J: Culture of Pancreatic Islet Cells,Cell & Tissue Culture: Laboratory Procedures Vol. 1,Doyle, A., Griffiths, J., and Newell, D., John Wiley and Sons, Ltd., 12B:11.1, 1995
大鼠	大鼠	Duct	Papain: 25 u/ml	DMEM /F-12	Githens, S: Pancreatic Duct Epithelial Cells,Cell & Tissue Culture: Laboratory Procedures Vol. 1,Doyle, A., Griffiths, J., and Newell, D., John Wiley and Sons, Ltd., 12B:12.1, 1995
大鼠	SD大鼠, 雄性, 40 - 100 克	Acinar	Hyaluronidase: 462 u/ml	Ham's F12	Hirschi, K., Kenny, S., Justice, J., Brannon, P.: Effects of Secretin And Caerulein On Enzymes Of Cultured Pancreatic Acinar Cells, In Vitro Cell Dev Biol 27, 660, 1991
大鼠	SD大鼠, 雄性, 100 - 150 克	Parotid acinar	Trypsin: 0.001%	F12 medium	Yeh, C., Mertz, P., Oliver, C., Baum, B., and Kousvelari, E.: Cellular Characteristics of Long-Term Cultured Rat Parotid Acinar Cells, In Vitro Cell Dev Biol 27, 707, 1991
大鼠	Wistar大鼠, 雄性	Parotid acinar	Trypsin: 0.02%	Solution B	Foskett, J., Roifman, C. and Wong, D.: Activation of Calcium Oscillations by Thapsigargin in Parotid Acinar Cells, J Biol Chem 266, 2778, 1991
大鼠	Wistar大鼠, 雄性	Parotid acinar	Trypsin: 0.02%	(See reference)
大鼠	Wistar大鼠, 雄性, 230 - 270 克	Islets	Collagenase: 0.1% - 0.2%	HBSS	Ohzato, H., Gotoh, M., Monden, M., Dono, K., Kanai, T., and Mori, T.: Improvement in the Islet Yield From a Cold-Preserved Pancreas by Pancreatic Ductal Collagenase Distention at the Time of Harvesting, Transplantation 51, 566, 1991
大鼠	(包括小鼠, 6 - 8 周大)	Islets	Collagenase: 0.1% - 0.2%	HBSS
大鼠	SD大鼠, 任一性別, 6-20 周大	Interlobular ducts	Papain: 25 u/ml	DMEM/Ham's F-12	Githens, S., Schexnayder, J., Desai, K., and Patke, C.: Rat Pancreatic Interlobular Duct Epithelium: Isolation and Culture in Collagen Gel, In Vitro Cell Dev Biol 25 (8), 679, 1989
大鼠	SD大鼠, 雄性, 150 - 200 克	Acinar	Soybean Trypsin Inhibitor: 0.01%	HEPES	Menozzi, D., Sato, S., Jensen, R., and Gardner, J.: Cyclic GMP Does Not Inhibit Protein Kinase C-Mediated Enzyme Secretion in Rat Pancreatic Acini, J Biol Chem 264, 995, 1989
大鼠	Wistar大鼠, 雄性, 250 - 350 克	Acinar, parotid	Hyaluronidase: 0.015%	Earle's MEM	Melvin, J., Kawaguchi, M., Baum, B., and Turner, R.: A Muscarinic Agonist-Stimulated Chloride Efflux Pathway is Associated With Fluid Secretion in Rat Parotid Acinar Cells,Biochem Biophys Res Commun 145, 754, 1987
大鼠	SD大鼠, 雄性, 50 - 125 克	Acinar	Trypsin: 0.01%	HBSS, CMF	Oliver, C., Waters, J., Tolbert, C., and Kleinman, H.: Growth of Exocrine Acinar Cells on a Reconstituted Basement Membrane Gel, In Vitro Cell Dev Biol 23, 465, 1987
大鼠	SD大鼠, 雄性, 50 - 125 克	Exorbital lacrimal, parotid, pancreas	Trypsin: 0.01%	HBSS, CMF
大鼠	Fischer-344大鼠, 任一性別, 120-150 克	Epithelial	Trypsin: 0.1%	Ham's F-12/HBSS	Tsao, M., and Duguid, W.: Establishment of Propagable Epithelial Cell Lines From Normal Adult Rat Pancreas, Exp Cell Res 168, 365, 1987
大鼠	Fischer-344大鼠, 任一性別, 120-150 克	Epithelial	Trypsin: 0.1%	(see reference)
大鼠	大鼠, 200-350 克	Islets	Collagenase Type 4: 0.2%	HBSS	Gotoh, M., Maki, T., Satomi, S., Porter, J., Bonner-Weir, S., O'Hara, C., and Monaco, A.: Reproducible High Yield of Rat Islets By Stationary In Vitro Digestion Following Pancreatic Ductal or Portal Venous Collagenase Injection, Transplantation 43 (5), 725, 1987
大鼠	SD大鼠, 雄性, 42 - 48 天, 175 - 200 克	Acinar, submandibular gland	Hyaluronidase: 0.1 %	HBSS, CF	Quissell, D., Redman, R., and Mark, M.: Short-Term Primary Culture of Acinar-Intercalated Duct Complexes From Rat Submandibular Glands, In Vitro Cell Dev Biol 22, 469, 1986
大鼠	暗條紋和劉易斯大鼠, 供血者, 成年的	Islets	Collagenase Type 1: 0.3%	Hank's solution	Sutton, R., Peters, M., McShane, P., Gray, D., and Morris, P.: An Improved Method for the Isolation of Islets of Langerhans From the Adult Rat Pancreas, Transplant Proc XVII (6), 1819, 1986
大鼠	SD大鼠, 雄性, 50-75 克	Acinar	Hyaluronidase: 0.1%	HBSS	Brannon, P., Orrison, B., and Kretchmer, N.: Primary Cultures of Rat Pancreatic Acinar Cells in Serum-Free Medium, In Vitro Cell Dev Biol 21 (1), 6, 1985
大鼠	SD大鼠, 雄性, 50-75 克	Acinar	Hyaluronidase: 0.1%	(see reference)
大鼠	SD大鼠, 125 - 350 克	Duct	Trypsin: 0.01%	HBSS	Githens, S., Holmquist, D., Whelan, J., and Ruby, J.: Ducts of the Rat Pancreas in Agarose Matrix Culture, In Vitro 16, 797, 1980
大鼠	大鼠	Exocrine	Hyaluronidase: 0.9%	Kreb's Ringer	Schulz, I., Heil, K., Kribben, A., Sachs, G., and Haase, W.: Isolation and Functional Characterization of Cells From the Exocrine Pancreas,Biology of Normal and Cancerous Exocrine Pancreatic Cells,Ribet, A., Pradayrol, L., and Susini, C., Elsevier, , 1980
大鼠	Wistar大鼠, 雄性	Islets	Collagenase Type 4: 1%	Medium 199	Katada, T., and Ui, M.: Enhanced Insulin Secretion and Cyclic Amp Accumulation in Pancreatic Islets Due to Activation of Native Calcium Ionophores, J Biol Chem 254 (2), 469, 1979
大鼠	SD大鼠, 雄性, 250 - 350 克	Acinar	Hyaluronidase: 0.18%	Kreb's Henseleit bicarbonate buffer	Williams, J., Korc, M., and Dormer, R.: Action of Secretagogues on a New Preparation of Functionally Intact, Isolated Pancreatic Acini, Am J Physiol 235, 517, 1978
大鼠	(包括小鼠, 小白鼠, 瑞士鼠, 雄性, 20 - 24 克)	Acinar	Hyaluronidase: 0.18%	Kreb's Henseleit bicarbonate buffer
大鼠	Wistar大鼠, 雄性, 300-350 克	Islets	Collagenase: 0.2%	Kreb's Ringer bicarbonate buffer	Wolters, G., Konijenendijk, W., and Bouman, P.: Effects of Fasting on Insulin Secretion, Islet Glucose Metabolism, and the Cyclic Adenosine 3'5'-Monophosphate Content of Rat Pancreatic Islets In Vitro, Diabetes 26 (6), 530, 1977
大鼠	Wistar大鼠, 雄性, 200 - 300 克	Islets	Collagenase Type 4: 0.5%	HBSS	Shibata, A., Ludvigsen, C., Naber, S., McKaneil M., and Lacy, P.: Standardization of a Digestion-Filtration Method for Isolation of Islets, Diabetes 25, 667, 1976
大鼠	大鼠	Exocrine	Hyaluronidase: 0.15%	Krebs	Kondo, S., and Schulz, I.: Calcium Ion Uptake in Isolated Pancreas Cells Induced by Secretagogues, Biochim Biophys Acta 419, 76, 1976
大鼠	Wistar大鼠-Lewis大鼠, 胎兒 18-20 妊娠日	Islets	Collagenase Type 4: 0.63%	EBSS	Braaten, J., Jarlfors, U., Smith, D., and Mintz, D.: Purification of Monolayer Cell Cultures of the Endocrine Pancreas, Tissue Cell 7 (4), 747, 1975
大鼠	(包括新生 3-12 天大)	Islets	Collagenase Type 4: 0.63%	(see reference)
大鼠	SD大鼠, 白話病變, 雄性, 150-200 克	Acinar, parotid	Trypsin: 0.01%	HBSS CMF	Mangos, J., McSherry, N., Butcher, F., Irwin, K., and Barber, T.: Dispersed Rat Parotid Acinar Cells (Morphological, Functional Character), Am J Physiol 229 (3), 560, 1975
			Collagenase: 40-50 u/ml
			Hyaluronidase: 0.10%
大鼠	大鼠, 新生的	Islets	Trypsin: 0.05%	Puck's saline buffered w/ EDTA 0.02%	Leonard, R., Lazarow, A., and Hegre, O.: Pancreatic Islet Transplantation in the Rat,Diabetes 22, 413, 1973
大鼠	大鼠, 雄性, 白話病變, 200-300 克	Islets	Collagenase: 0.5%	Hanks solution	Lacy, P., Walker, M., and Fink, J.: Perifusion of Isolated Rat Islets in Vitro, Diabetes 21 (10), 987, 1972
大鼠	Lewis大鼠, 近親繁殖的	Islets	Collagenase: 0.5%	HBSS	Ballinger, W., and Lacy, P.: Transplantation of Intact Pancreatic Islets in Rats, Surgery 72 (2), 175, 1972
大鼠	Wistar大鼠, 白話病變, 雄性, 400 - 500 克	Islets	Collagenase Type 4: 1.0% - 1.2%	HBSS	Lacy, P., and Kostianovsky, M.: Method for the Isolation of Intact Islets of Langerhans from the Rat Pancreas, Diabetes 16, 35, 1967
大鼠	大鼠	Ascites hepatoma	Trypsin: 0.1%	Phosphate buffer (see reference)	Essner, E.: Experiments on an Ascites Hepatoma. I. Enzymatic Digestion and Alkaline Degradation of the Cementing Substance and Separation of Cells, in Tumor Islands, Exp Cell Res 7, 430, 1954

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