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干细胞组织消化手册

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2016/2/16 16:02:27

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Worthington组织消化指南

引言

组织消化酶主要用于组织培养和细胞生物学研究的原代细胞的分离和收集。尽管这些消化酶已经普遍应用多年,但是对其组织消化和细胞收集的作用机制却没有全面的理解。  由此导致的结果便是选择一种消化方法大多是随机选择或基于过去的经验,而不是在*理解消化原理的基础上选择的消化方案,以及如何根据具体实验目的修正消化方案以获得更好的实验结果。

细胞分离程序的目的是zui大程度的获得功能活性的单个细胞,而影响结果的原因有很多,包括(不限于)以下:

1、组织类型

2、物种来源

3、动物年龄

4、遗传改造情况(如knockouts

5、所用的消化培养液

6、所用的酶

7、消化酶中的杂质

8、所用消化酶的工作浓度

9、温度

10孵育时间

通常前四个原因没办法选择,而剩下的原因一般由经验决定以获取zui适实验结果。

通过查阅文献来选择*组织消化酶和zui适消化条件的研究人员通常被互相矛盾的实验数据搞得一头雾水。这些巨大差异来源于组织胞外基质的复杂而动态的特性,以及过去常用的粗制、未提纯的组织分离酶。另外,组织胞外基质由各种蛋白、糖蛋白、脂类和糖脂组成,这些成分的组成比例在不同物种、不同组织、甚至不同发育(年龄)之间存在明显的差异,这些因素都极大的影响了消化方案的制定。目前通常使用的粗制酶,如链霉蛋白酶、NF1:250、以及含有几种其他不同浓度蛋白酶的胶原酶、以及各种多糖酶、核酶和脂酶。

本指南总结了目前常用组织消化酶的zui适消化和细胞收集知识和方法,描述了标准的实验步骤, 便于制定一个*的细胞分离方案,同时列举了一些相关的参考文献。

下表列出多种组织的适宜消化酶(含参考文献,以组织类型和物种分类,详见链接)

脂肪组织

肾上腺组织

骨组织

脑组织

软骨组织

结肠组织

内皮组织

上皮组织

心脏

淋巴结

乳腺

其它组织

其他组织

肌组织

神经组织

胰腺

腮腺

垂体

前列腺

生殖

动物鳞片

皮肤

脾脏

干细胞

胸腺

甲状腺/甲状旁腺

扁桃体

肿瘤

 

 

 

注:使用Worthington公司消化酶的文献不限于上表链接中所列,一般而言,Worthington公司的组织消化酶可用于大多组织类型的消化。

 

干细胞组织消化手册

 

物种详细来源细胞名称消化用酶培养基参考文献
Adipose derived stem cellsCollagenase Type 1: 0.1%DMEMTandon, N., Goh, B., Marsano, A., Chao, P., Montouri-Sorrentino, C., Gimble, J. and Vunjak-Novakovic, G.: Alignment and Elongation of Human Adipose-Derived Stem Cells in Response to Direct-Current Electrical Stimulation., Conf Proc IEEE Eng Med Biol Soc 2009, 6517-21, 2009
Amniotic membrane MSCTrypsin: 0.25%LG-DMEMPark, S., Seo, M., Kim, H. and Kang, K.: Isolation and Characterization of Canine Amniotic Membrane-Derived Multipotent Stem Cells., PLoS ONE 7, e44693, 2012
Collagenase Type 1: 0.2%
犬, 20-25 公斤Adipose stem cellsCollagenase: see referenceMedia-199Fischer, L., McIlhenny, S., Tulenko, T., Golesorkhi, N., Zhang, P., Larson, R., Lombardi, J., Shapiro, I. and DiMuzio, P.: Endothelial Differentiation of Adipose-Derived Stem Cells: Effects of Endothelial Cell Growth Supplement and Shear Force., J Surg Res 152, 157, 2009
Adipose derived stem cellsCollagenase Type 1: 0.1%PBSVidal, M., Robinson, S., Lopez, M., Paulsen, D., Borkhsenious, O., Johnson, J., Moore, R. and Gimble, J.: Comparison of Chondrogenic Potential in Equine Mesenchymal Stromal Cells Derived from Adipose Tissue and Bone Marrow., Vet Surg 37, 713, 2008
马, 1-5 岁Adipose derived stem cellsCollagenase Type 1: 0.1%PBSVidal, M., Kilroy, G., Lopez, M., Johnson, J., Moore, R. and Gimble, J.: Characterization of Equine Adipose Tissue-Derived Stromal Cells: Adipogenic and Osteogenic Capacity and Comparison with Bone Marrow-Derived Mesenchymal Stromal Cells., Vet Surg 36, 613, 2007
人类人类Mesenchymal Stromal CellsCollagenase Type 4: 0.2%PBSChoudhery,M, Badowski, M, Muise, A and Harris, D: Effect of Mild Heat Stress on the Proliferative and Differentiative Ability of Human Mesenchymal Stromal Cells., Cytotherapy17, 359-68, 2015
人类人类Adipose derived stem cellsCollagenase Type 2: 0.1%HBSSSatish, L., Krill-Burger, J., Gallo, P., Etages, S., Liu, F., Philips, B., Ravuri, S., Marra, K., LaFramboise, W., Kathju, S. and Rubin, J.: Expression Analysis of Human Adipose-Derived Stem Cells During In Vitro Differentiation to an Adipocyte Lineage., BMC Med Genomics 8, 41, 2015
人类人类Dental mesenchymal stemCollagenase Type 1: 0.3%DMEMWu, Z., Wang, J., Dong, R., Wang, L., Fan, Z., Liu, D. and Wang, S.: Depletion of MEIS2 Inhibits Osteogenic Differentiation Potential of Human Dental Stem Cells., Int J Clin Exp Med8, 7220-30, 2015
Neutral Protease: 0.4%
人类人类Decidua Mesenchymal StemTrypsin: 0.25%PBSKusuma, G., Menicanin, D., Gronthos, S., Manuelpillai, U., Abumaree, M., Pertile, M., Brennecke, S. and Kalionis, B.: Ectopic Bone Formation by Mesenchymal Stem Cells Derived from Human Term Placenta and the Decidua., PLoS ONE 10, e0141246, 2015
Collagenase Type 1: 1.0%
Deoxyribonuclease I: 0.005%
人类人类, 成年人Adipose derived mesenchymal stemCollagenase Type 2: 0.1%see referenceAl-Saqi, S., Saliem, M., Asikainen, S., Quezada, H., Ekblad, A., Hovatta, O., Le Blanc, K., Jonasson, A. and Gotherstrom, C.: Defined Serum-Free Media for In Vitro Expansion of Adipose-Derived Mesenchymal Stem Cells., Cytotherapy 16, 915, 2014
人类人类, 成年人Adipose derived stem cellsCollagenase Type 1: 0.15%DMEMKoellensperger, E., Bollinger, N., Dexheimer, V., Gramley, F., Germann, G. and Leimer, U.: Choosing the Right Type of Serum for Different Applications of Human Adipose Tissue-Derived Stem Cells: Influence on Proliferation and Differentiation Abilities., Cytotherapy 16, 789, 2014
人类人类Umbilical mesenchymal stemCollagenase Type 2: 0.1%DMEMHang, H., Yu, Y., Wu, N., Huang, Q., Xia, Q. and Bian, J.: Induction of Highly Functional Hepatocytes from Human Umbilical Cord Mesenchymal Stem Cells by HNF4伪 Transduction.,PLoS ONE 9, e104133, 2014
Trypsin: 0.25%
人类人类Cardiac stemCollagenase Type 2: 0.25%Joklik modified Eagle抯Avolio, E., Gianfranceschi, G., Cesselli, D., Caragnano, A., Athanasakis, E., Katare, R., Meloni, M., Palma, A., Barchiesi, A., Vascotto, C., Toffoletto, B., Mazzega, E., Finato, N., Aresu, G. and Livi, U.: Ex vivo Molecular Rejuvenation Improves the Therapeutic Activity of Senescent Human Cardiac Stem Cells in a Mouse Model of Myocardial Infarction., Stem Cells 32, 2373, 2014
人类人类Adipose stromal stemCollagenase animal free: 200 u/mlDMEM/Hams F-12Carvalho, P., Gimble, J., Dias, I., Gomes, M. and Reis, R.: Xenofree Enzymatic Products for the Isolation of Human Adipose-Derived Stromal/Stem Cells., Tiss Eng 19, 473-8, 2013
人类人类, 成年人Epidermal stem cellsNeutral Protease: 2 u/mlDMEMNagel, S., Rohr, F., Weber, C., Kier, J., Siemers, F., Kruse, C., Danner, S., Brandenburger, M. and Matthiessen, A.: Multipotent Nestin-Positive Stem Cells Reside in the Stroma of Human Eccrine and Apocrine Sweat Glands and can be Propagated Robustly In Vitro., PLoS ONE 8, e78365, 2013
Trypsin: 2.5%
人类人类Dental pulp stemCollagenase Type 1: 0.3%DMEM/F12Bonnamain, V., Thinard, R., Sergent-Tanguy, S., Huet, P., Bienvenu, G., Naveilhan, P., Farges, J. and Alliot-Licht, B.: Human Dental Pulp Stem Cells Cultured in Serum-Free Supplemented Medium., Front Physiol 4, 357, 2013
Neutral Protease: 0.4%
人类人类Umbilical Cord Mesenchymal StemCollagenase: 0.1-0.2%DMEMHan, Y., Tao, R., Sun, T., Chai, J., Xu, G. and Liu, J.: Optimization of Human Umbilical Cord Mesenchymal Stem Cell Isolation and Culture Methods., Cytotechnology 65, 819, 2013
Trypsin: 0.1%
人类人类Dental pulp derived stem cellCollagenase Type 1: 0.3%DMEMSakai, K., Yamamoto, A., Matsubara, K., Nakamura, S., Naruse, M., Yamagata, M., Sakamoto, K., Tauchi, R., Wakao, N., Imagama, S., Hibi, H., Kadomatsu, K., Ishiguro, N. and Ueda, M: Human Dental Pulp-Derived Stem Cells Promote Locomotor Recovery After Complete Transection of the Rat Spinal Cord by Multiple Neuro-Regenerative Mechanisms., J Clin Invest 122, 80, 2012
Neutral Protease: 0.4%
人类人类Adipose-derived stem cellsCollagenase Type 1: 0.1%PBSCervelli, V., Scioli, M., Gentile, P., Doldo, E., Bonanno, E., Spagnoli, L. and Orlandi, A.:Plaet-Rich Plasma Greatly Potentiates Insulin-Induced Adipogenic Differentiation of Human Adipose-Derived Stem Cells Through a Serine/Threonine Kinase Akt-dependent Mechanism and Promotes Clinical Fat Graft Maintenance., Stem Cells Transl Med 1, 206-20, 2012
人类人类Mesenchymal stemCollagenase Type 1: 0.4%DMEM/F12Shalini, V., Pratheep, S., Muhammad, A., Sharmili, V., Elizabeth, G. and Rajesh, R.: Generation and Characterisation of Human Mesenchymal Stem Cells Derived From Umbilical Cord and Placenta, Regenerative Research 1, 48, 2012
Deoxyribonuclease I: 0.01%
人类人类Wharton jelly mesenchymal stemCollagenase Type 2: 0.1%DMEMSalehinejad, P., Alitheen, N., Ali, A., Omar, A., Mohit, M., Janzamin, E., Samani, F., Torshizi, Z. and Nematollahi-Mahani, S.: Comparison of Different Methods for the Isolation of Mesenchymal Stem Cells from Human Umbilical Cord Wharton's Jelly., In Vitro Cell Dev Biol Anim 48, 75, 2012
Trypsin: 0.25%
人类人类Placental Mesenchymal StemCollagenase Type 2: 0.4%DMEMVellasamy, S., Sandrasaigaran, P., Vidyadaran, S., George, E. and Ramasamy, R.:Isolation and Characterisation of Mesenchymal Stem cells Derived from Human Placenta Tissue., World J Stem Cells 4, 53-61, 2012
Deoxyribonuclease I: 0.01%
人类人类, 男性 40-60 yearsAdipose derived stem cellsCollagenase: 0.25%PBSBlasi, A., Martino, C., Balducci, L., Saldarelli, M., Soleti, A., Navone, S., Canzi, L., Cristini, S., Invernici, G., Parati, E. and Alessandri, G.: Dermal Fibroblasts Display Similar Phenotypic and Differentiation Capacity to Fat-Derived Mesenchymal Stem Cells, but Differ in Anti-Inflammatory and Angiogenic Potential, Vasc Cell 3, 5, 2011
Deoxyribonuclease I: 0.002%
人类人类Umbilical cord stromal stemCollagenase Type 4: 0.08%DMEMFarias, V., Linares-Fernandez, J., Penalver, J., Paya Colmenero, J., Ferron, G., Duran, E., Fernandez, R., Olivares, E., O'Valle, F., Puertas, A., Oliver, F. and Ruiz de Almodovar, J.:Human Umbilical Cord Stromal Stem Cell Express CD10 and Exert Contractile Properties.,Placenta 32, 86, 2011
Neutral Protease: 0.138%
Hyaluronidase: 0.02%
人类人类Vascular wall-resident multipotent stem cellsCollagenase Type 2: 0.2%see referenceKlein, D., Weisshardt, P., Kleff, V., Jastrow, H., Jakob, H., and Ergun, S.: Vascular Wall-Resident CD44+ Multipotent Stem Cells Vive Rise to Pericytes and Smooth Muscle Cells and Contribute to New Vessel Maturation., PLoS ONE 6, e20540, 2011
Elastase: 5 u/ml
人类人类Adipose derived stemCollagenase Type 1: 0.1%DMEM/Ham's F-12Yu, G. Floyd, ZE., Wu, X., Halvorsen, Y. and Gimble, J.: Isolation of Human Adipose-Derived Stem Cells from Lipoaspirates., Methods Mol Biol 702, 17-27, 2011
人类人类Glioma stem cellsPDS kit: per instructionsneurobasal mediumHjelmeland, A., Wu, Q., Wickman, S., Eyler, C., Heddleston, J., Shi, Q., Lathia, J., Macswords, J., Lee, J., McLendon, R. and Rich, J.: Targeting A20 Decreases Glioma Stem Cell Survival and Tumor Growth., PLoS Biol 8, e1000319, 2010
人类人类Dental pulp and apical papilla stem cellsCollagenase Type 1: 0.3%MEMHuang, G., Yamaza, T., Shea, L., Djouad, F., Kuhn, N., Tuan, R. and Shi, S.:Stem/Progenitor Cell-Mediated De Novo Regeneration of Dental Pulp with Newly Deposited Continuous Layer of Dentin in an In Vivo Model., Tissue Eng Part A 16, 605, 2010
Neutral Protease: 0.4%
人类人类Corneal stromal stemNeutral Protease: 1.2 u/mlDMEMDu, Y., Roh, D., Funderburgh, M., Mann, M., Marra, K., Rubin, J., Li, X. and Funderburgh, J.: Adipose-Derived Stem Cells Differentiate to Keratocytes In Vitro., Mol Vis 16, 2680, 2010
Collagenase: 0.1%
人类人类Adipose tissue-derived stemCollagenase Type 2: 1.0%DMEM/F12Tan, H., DeFail, A., Rubin, J., Chu, C. and Marra, K.: Novel Multiarm PEG-Based Hydrogels for Tissue Engineering., J Biomed Mater Res 92, 979, 2010
人类人类Umbilical cord mesenchymalHyaluronidase: 0.05%DMEMZeddou, M., Briquet, A., Relic, B., Josse, C., Malaise, M., Gothot, A., Lechanteur, C. and Beguin, Y.: The Umbilical Cord Matrix is a Better Source of Mesenchymal Stem Cells (MSC) Than the Umbilical Cord Blood., Cell Biol Int 34, 693-701, 2010
Collagenase: 0.08%
人类人类Adipose stem cellsCollagenase Type 1: 0.1%DMEMHareendran, S., Sathishkumar, S., Abbas, S., Mackay, A. and Rajan, P.: A Novel Composition for the Culture of Human Adipose Stem Cells which Includes Complement C3.,Cytotechnology 62, 389-402, 2010
人类人类, 40-65 yearAdult human adipose stem cellsCollagenase Type 2: 0.075%DMEMSun, N., Panetta, N., Gupta, D., Wilson, K., Lee, A., Jia, F., Hu, S., Cherry, A., Robbins, R., Longaker, M. and Wu, J.: Feeder-Free Derivation of Induced Pluripotent Stem Cells from Adult Human Adipose Stem Cells., Proc Natl Acad Sci U S A 106, 15720, 2009
人类人类Filum terminale neural progenitorTrypsin: see referenceDMEM/F-12Varghese, M., Olstorn, H., Berg-Johnsen, J., Moe, M., Murrell, W. and Langmoen, I.:Isolation of Human Multipotent Neural Progenitors from Adult Filum Terminale., Stem Cells Dev 18, 603, 2009
人类人类skin and keloid progenitor cellsCollagenase Type 1: 0.4%PBSZhang, Q., Yamaza, T., Kelly, AP, Shi, S., Wang, S., Brown, J., Wang, L., French, S., Shi, S. and Le, A.: Tumor-like Stem Cells Derived from Human Keloid are Governed by the Inflammatory Niche Driven by IL-17/IL-6 Axis., PLoS ONE 4, e7798, 2009
Neutral Protease: 0.3%
人类人类, 男性Spermatogonial stem cellsCollagenase: 1%DMEMKossack, N., Meneses, J., Shefi, S., Nguyen, H., Chavez, S., Nicholas, C., Gromoll, J., Turek, P. and Reijo-Pera, R.: Isolation and Characterization of Pluripotent Human Spermatogonial Stem Cell-Derived Cells., Stem Cells , , 2008
Deoxyribonuclease I: 0.22%
Trypsin: 0.4%
人类人类Muscle derived multiprogenitor cellsCollagenase Type 2: 0.05%DMEMNesti, L., Jackson, W., Shanti, R., Koehler, S., Aragon, A., Bailey, J., Sracic, M., Freedman, B., Giuliani, J. and Tuan, R.: Differentiation Potential of Multipotent Progenitor Cells Derived from War-Traumatized Muscle Tissue., J Bone Joint Surg Am 90, 2390, 2008
人类人类Adipose derived stem cellsCollagenase Type 1: 0.075%DMEMJeong, J.: Adipose Stem Cells as a Clinically Available and Effective Source of Adult Stem Cell Therapy, Int J Stem Cells 1, 43, 2008
人类人类Hepatic stem cells and heptoblastsCollagenase Type 4: 0.014-0.06%variousWauthier, E., Schmelzer, E., Turner, W., Zhang, L., LeCluyse, E., Ruiz, J., Turner, R., Furth, M., Kubota, H., Lozoya, O., Barbier, C., McClelland, R., Yao, H., Moss, N., Bruce, A., Ludlow, J. and Reid, L.: Hepatic Stem Cells and Hepatoblasts: Identification, Isolation, and Ex Vivo Maintenance., Methods Cell Biol 86, 137, 2008
人类人类Umbilical mesenchymal stemCollagenase: 0.1%DMEMSecco, M., Zucconi, E., Vieira, N., Fogaca, L.,Cerqueira, A., Carvalho, M., Jazedje, T., Okamoto, O., Muotri, A. and Zatz, M.: Multipotent Stem Cells from Umbilical Cord: Cord is Richer Than Blood!, Stem Cells 26, 146, 2008
人类人类Stem CellsCollagenase: 280 u/mlD-PBSPilgaard, L., Lund, P., Rasmussen, J., Fink, T. and Zachar, V.: Comparative Analysis of Highly Defined Proteases for the Isolation of Adipose Tissue-Derived Stem Cells., Regen Med3, 705-15, 2008
人类人类Bone marrow derived MSCTrypsin: 0.05%DMEMWelter, J., Solchaga, L.and Penick, K.: Simplification of Aggregate Culture of Human Mesenchymal Stem Cells as a Chondrogenic Screening Assay, Biotechniques 42, 732, 2007
Papain: 0.0025%
人类人类, 8-12 yearTendon stem/progenitorCollagenase Type 1: 0.3%DMEMBi Yanming, Ehirchiou Driss, Kilts Tina M, Inkson Colette A, Embree Mildred C, Sonoyama Wataru, Li Li, Leet Arabella I, Seo Byoung-Moo, Zhang Li, Shi Songtao, Young Marian F:Identification of tendon stem/progenitor cells and the role of the extracellular matrix in their niche, Nat Med 13, 1219-27, 2007
Neutral Protease: 0.4%
人类人类, 男性Adipose derived adult stem cellsCollagenase Type 1: 0.1%DMEMLei, L., Liao, W., Sheng, P., Fu, M., He, A. and Huang, G.: Biological Character of Human Adipose-Derived Adult Stem Cells and Influence of Donor Age on Cell Replication in Culture.,Sci China C Life Sci 50, 320, 2007
人类人类Pancreatic cancer stem cellsCollagenase Type 4: 200 u/mlmedium 199Li, 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
人类人类, 成年人Adipose-derived adult stemCollagenase Type 1: 0.1%DMEM/F-12 Ham'sMitchell James B, McIntosh Kevin, Zvonic Sanjin, Garrett Sara, Floyd Z Elizabeth, Kloster Amy, Di Halvorsen Yuan, Storms Robert W, Goh Brian, Kilroy Gail, Wu Xiying, Gimble Jeffrey M: Immunophenotype of human adipose-derived cells: temporal changes in stromal-associated and stem cell-associated markers, Stem Cells 24, 376-85, 2006
人类人类Mesenchymal stemCollagenase Type 1: 0.075%DMEMKern Susanne, Eichler Hermann, Stoeve Johannes, Kluter Harald, Bieback Karen:Comparative analysis of mesenchymal stem cells from bone marrow, umbilical cord blood, or adipose tissue, Stem Cells 24, 1294-301, 2006
人类人类Placental mesenchymal stemCollagenase Type 2: 270 u/mlMEMPortmann-Lanz CBettina, Schoeberlein Andreina, Huber Alexander, Sager Ruth, 男性k Antoine, Holzgreve Wolfgang, Surbek DanielV: Placental mesenchymal stem cells as potential autologous graft for pre- and perinatal neuroregeneration, Am J Obstet Gynecol194, 664-73, 2006
Neutral Protease: 2.4 u/ml
人类人类Stromal stem cellsCollagenase Type 1: 0.2%HBSSBoquest Andrew C, Shahdadfar Aboulghassem, Brinchmann Jan E, Collas Philippe:Isolation of stromal stem cells from human adipose tissue, Methods Mol Biol 325, 35-46, 2006
人类人类, 女性Adipose derived adult stem cellsCollagenase Type 1: 0.1%DMEM/F-12Guilak, F., Lott, K., Awad, H., Cao, Q., Hicok, K., Fermor, B. and Gimble, J.: Clonal Analysis of the Differentiation Potential of Human Adipose-Derived Adult Stem Cells., J Cell Physiol206, 229, 2006
人类人类Adult stem cellsCollagenase Type 1: 0.1%PBSDevireddy, R., Thirumala, S. and Gimble, J.: Cellular Response of Adipose Derived Passage-4 Adult Stem Cells to Freezing Stress., J Biomech Eng 127, 1081, 2005
人类人类, 女性Adipose derived adult stem cellsCollagenase Type 1: 0.1%DMEM-Ham's F-12Aust, L., Devlin, B., Foster, S., Halvorsen, Y., Hicok, K., du Laney, T., Sen, A., Willingmyre, G. and Gimble, J.: Yield of Human Adipose-Derived Adult Stem Cells from Liposuction Aspirates., Cytometry 6, 7-14, 2004
人类人类, 成年人, 男性Human epidermal keratinocyte stem cellsNeutral Protease: 0.5%DMEMPapini S, Cecchetti D, Campani D, Fitzgerald W, Grivel J, Chen S, Margolis L, Revolla R.: Isolation and Clonal Analysis of Human Epidermal Keratinocyte Stem Cells in Long-term Culture, Stem Cells 21, 481, 2003
人类人类Umbilical vein mesenchymal stem cellsCollagenase: 1%PBSCovas DT, Siufi JL C, Silva AR L, Orellana MD: Isolation and culture of umbilical vein mesenchymal stem cells, Braz J Med Biol Res 36, 1179-83, 2003
人类人类, 7-8 岁Stem cells Human exfoliated deciduous teethCollagenase Type 1: 0.3%see referenceMiura, M., Gronthos, S, Zhao, M, Lu, B, Fisher, L, Robey, P, and Shi, S.: SHED: Stem Cells from Human Exfoliated Deciduous Teeth, Proc Natl Acad Sci U S A 100, 5807, 2003
Neutral Protease: 0.4%
人类人类, 胎儿Epithelial progenitorCollagenase: 0.03%DMEMMalhi, H., Irani, A., Gagandeep, S. and Gupta, S.: Isolation of Human Progenitor Liver Epithelial Cells with Extensive Replication Capacity and Differentiation into Mature Hepatocytes., J Cell Sci 115, 2679, 2002
人类人类, 成年人Human skin mast cellsCollagenase Type 2: 0.15%HBSSKambe, N, Kambe, M, Kochan, J, and Schwartz ,L.: Human Skin-derived Mast Cells Can Proliferate While Retaining Their Characteristic Functional and Protease Phenotypes, Blood 97, 2045, 2001
Hyaluronidase: 0.07%
Deoxyribonuclease I: 0.03%
人类人类Central nervous system stemCollagenase: 0.1%HBSSUchida, N., Buck, D., He, D., Reitsma, M., Masek, M., Phan, T., Tsukamoto, A., Gage, F. and Weissman, I.: Direct Isolation of Human Central Nervous System Stem Cells., Proc Natl Acad Sci U S A 97, 14720, 2000
Hyaluronidase: 0.1%
人类人类Embryonic stemNeutral Protease: 1%DMEMThomson JA, Itskovitz-Eldor J, Shapiro SS, Waknitz MA, Swiergiel JJ, Marshall VS, Jones JM: Embryonic stem cell lines derived from human blastocysts, Science 282, 1145-7, 1998
Collagenase Type 4: 0.1%
人类人类Muscle-derived stem cellsTrypsin: 0.25%DMEM/F12Alessandri Giulio, Pagano Stefano, Bez Alessandra, Benetti Anna, Pozzi Stefano, Iannolo Gioacchin, Baronio Manuela, Invernici Gloria, Caruso Arnaldo, Muneretto Claudio, Bisleri Gianluigi, Parati Eugenio: Isolation and culture of human muscle-derived stem cells able to differentiate into myogenic and neurogenic cell lineages, Lancet 364, 1872-83,
Embryonic stemCollagenase Type 4: 0.08%DMEMChen, S., Revolla, R., Papini, S., Michelini, M., Fitzgerald, W., Zimmerberg, J.,and Margolis, L.: Multilineage Differentiation of Rhesus Monkey Embryonic Stem cells in Three-dimensional Culture Systems, Stem Cells 21(3), 281, 2003
Baboons, 1岁, 7岁, 14 岁Primate spermatogonialCollagenase Type 2: 0.1%DMEMNagano, M., McCarrey, J., and Brinster, R.: Primate Spermatogonial Stem Cells Colonize Mouse Testes, Biol Reprod 64, 1409, 2001
Trypsin: 0.05%
Deoxyribonuclease I: 0.1%
小鼠小鼠, 3-4 周Cardiac progenitorCollagenase Type 1: 0.1%DMEMWang, H., Chen, H., Feng, B., Wang, X., He, X., Hu, R., Yin, M., Wang, W., Fu, W. and Xu, Z.: Isolation and Characterization of a Sca-1+/CD31- Progenitor Cell Lineage Derived From Mouse Heart Tissue., BMC Biotechnol 14, 75, 2014
小鼠小鼠, P4Enteric neural crest progenitorsNeutral Protease: 0.5%DMEM/F12Hotta, R., Stamp, L., Foong, J., McConnell, S., Bergner, A., Anderson, R., Enomoto, H., Newgreen, D., Obermayr, F., Furness, J. and Young, H.: Transplanted Progenitors Generate Functional Enteric Neurons in the Postnatal Colon., J Clin Invest 123, 1182, 2013
Collagenase animal free: 0.05%
小鼠小鼠Epithelial StemTrypsin: 0.25%HBSSJensen, U., Ghazizadeh, S. and Owens, D.: Isolation and Characterization of Cutaneous Epithelial Stem Cells., Methods Mol Biol 989, 61-9, 2013
Collagenase Type 1: 0.4%
小鼠小鼠, 10 周Synovial mesenchymalCollagenase: 0.1%DMEMFutami, I., Ishijima, M., Kaneko, H., Tsuji, K., Ichikawa-Tomikawa, N., Sadatsuki, R., Muneta, T., Arikawa-Hirasawa, E., Sekiya, I. and Kaneko, K.: Isolation and Characterization of Multipotential Mesenchymal Cells from the Mouse Synovium., PLoS ONE7, e45517, 2012
Deoxyribonuclease I: 0.005%
小鼠小鼠, 6-8 周Neural stem cellsPapain: 0.01%DMEM/F12Bracko, O., Singer, T., Aigner, S., Knobloch, M., Winner, B., Ray, J., Clemenson, G., Suh, H., Couillard-Despres, S., Aigner, L., Gage, F. and Jessberger, S.: Gene Expression Profiling of Neural Stem Cells and their Neuronal Progeny Reveals IGF2 as a Regulator of Adult Hippocampal Neurogenesis., J Neurosci 32, 3376-87, 2012
Neutral Protease: 0.1%
Deoxyribonuclease I: 0.01%
小鼠小鼠, 8-10 周Adipose derived stemCollagenase Type 1: 0.025%HBSSSugii, S., Kida, Y., Berggren, W. and Evans, R.: Feeder-Dependent and Feeder-Independent iPS Cell Derivation from Human and Mouse Adipose Stem Cells., Nat Protoc 6, 346, 2011
小鼠小鼠Spleen, bone marrow endothelialCollagenase Type 4: 0.3-1.0%PBSShi, C., Jia, T., Mendez-Ferrer, S., Hohl, T., Serbina, N., Lipuma, L., Leiner, I., Li, M., Frenette, P. and Pamer, E.: Bone Marrow Mesenchymal Stem and Progenitor Cells Induce Monocyte Emigration in Response to Circulating Toll-Like Receptor Ligands., Immunity 34, 590, 2011
Deoxyribonuclease I: 20 u/ml
小鼠小鼠, 8-10 周Lung mesenchymal stemCollagenase Type 2: 0.2%HBSSChow, K., Jun, D., Helm, K., Wagner, D. and Majka, S.: Isolation & Characterization of Hoechst(low) CD45(negative) Mouse Lung Mesenchymal Stem Cells., J Vis Exp 56, e3159, 2011
小鼠小鼠, 新生的Dental Pulp Stem CellsCollagenase Type 4: 0.2%PBSJanebodin, K., Horst, O., Ieronimakis, N., Balasundaram, G., Reesukumal, K., Pratumvinit, B. and Reyes, M.: Isolation and Characterization of Neural Crest-Derived Stem Cells from Dental Pulp of Neonatal Mice., PLoS ONE 6, e27526, 2011
Neutral Protease: 1.2 u/ml
小鼠小鼠Stem and progenitorCollagenase Type 2: 0.2%HBSSHan, J., Koh, Y., Moon, H., Ryoo, H., Cho, C., Kim, I. and Koh, G.: Adipose Tissue is an Extramedullary Reservoir for Functional Hematopoietic Stem and Progenitor Cells., Blood 115, 957, 2010
小鼠小鼠, 6-8 周Bone marrow mesenchymal stemCollagenase Type 1: 0.25%RPMI 1640Xu, S., De Becker, A., Van Camp, B., Vanderkerken, K. and Van Riet, I.: An Improved Harvest and In Vitro Expansion Protocol for Murine Bone Marrow-Derived Mesenchymal Stem Cells., J Biomed Biotechnol 2010, 105940, 2010
小鼠小鼠, 成年人Adult neural stemTrypsin: 0.05%DMEMDeleyrolle, L. and Reynolds, B.: Isolation, Expansion, and Differentiation of Adult Mammalian Neural Stem and Progenitor Cells using the Neurosphere Assay., Methods Mol Biol 549, 91, 2009
小鼠小鼠, 6-8 周Prostate epithelial/stemCollagenase Type 2: 0.5%HBSSBurger, P., Gupta, R., Xiong, X., Ontiveros, C., Salm, S., Moscali, D. and Wilson, E.:High Aldehyde Dehydrogenase Activity: A Novel Functional Marker of Murine Prostate Stem/Progenitor Cells., Stem Cells 27, 2220-8, 2009
Trypsin: 0.05%
小鼠小鼠Adipose tissue-derived stemCollagenase Type 1: 0.075%Modified EaglesCho, K., Park, H., Park, H., Jung, J., Jeon, S., Kim, Y. and Roh, H.: IFATS Collection: Immunomodulatory Effects of Adipose Tissue-Derived Stem Cells in an Allergic Rhinitis Mouse Model., Stem Cells 27, 259-65, 2009
小鼠小鼠Germ cellsCollagenase Type 1: 100 u/mlHBSSBreault, D., Min, I., Carlone, D., Farilla, L., Ambruzs, D., Henderson, D., Algra, S., Montgomery, R., Wagers, A. and Hole, N.: Generation of mTert-GFP Mice as a Model to Identify and Study Tissue Progenitor Cells., Proc Natl Acad Sci U S A 105, 10420, 2008
小鼠小鼠Neural progenitor cellPapain: 10 ul/mlPBSHutton, S. and Pevny, L.: Isolation, Culture, and Differentiation of Progenitor Cells from the Central Nervous System., Cold Spring Harb. Protoc. 11, 5077, 2008
小鼠小鼠, 3-6 周Ear mesenchymal stemCollagenase Type 1: 0.2%DMEM/F12Staszkiewicz, J., Gimble, J., Manuel, J. and Gawronska-Kozak, B.: IFATS Collection: Stem Cell Antigen-1-Positive Ear Mesenchymal Stem Cells Display Enhanced Adipogenic Potential.,Stem Cells 26, 2666, 2008
小鼠小鼠, 胚胎HES-BC cellsTrypsin: 0.05%DMEMLu Shi-Jiang, Feng Qiang, Caballero Sergio, Chen Yu, Moore Malcolm A, Grant Maria B, Lanza Robert: Generation of functional hemangioblasts from human embryonic stem cells,Nat Methods 4, 501-9, 2007
小鼠小鼠, 6-8 周Tendon stem/progenitorCollagenase Type 1: 0.3%DMEMBi Yanming, Ehirchiou Driss, Kilts Tina M, Inkson Colette A, Embree Mildred C, Sonoyama Wataru, Li Li, Leet Arabella I, Seo Byoung-Moo, Zhang Li, Shi Songtao, Young Marian F:Identification of tendon stem/progenitor cells and the role of the extracellular matrix in their niche, Nat Med 13, 1219-27, 2007
Neutral Protease: 0.4%
小鼠小鼠Liver epithelial progenitor cellsCollagenase Type 4: 0.1%DMEMLi Wen-Lin, Su Juan, Yao Yu-Cheng, Tao Xin-Rong, Yan Yong-Bi, Yu Hong-Yu, Wang Xin-Min, Li Jian-Xiu, Yang Yong-Ji, Lau Joseph T Y, Hu Yi-Ping: Isolation and characterization of bipotent liver progenitor cells from adult mouse, Stem Cells 24, 322-32, 2006
Deoxyribonuclease I: 0.05%
小鼠小鼠, 6 周Adipose mesenchymal stemCollagenase: 0.2%PBSDi Rocco Giuliana, Iachininoto Maria Grazia, Tritarelli Alessandra, Straino Stefania, Zacheo Antonella, Germani Antonia, Crea Filippo, Capogrossi Maurizio C: Myogenic potential of adipose-tissue-derived cells, J Cell Sci 119, 2945-52, 2006
小鼠小鼠Neural stem cellsPapain: see referenceDMEM/F-12Meletis, K., Wirta, V., Hede, S., Nister, M., Lundeberg, J. abd Frisen, J.: p53 Suppresses the Self-Renewal of Adult Neural Stem Cells., Development 133, 363, 2006
小鼠小鼠, 7天Cerebellar stem cellsPapain: 10u/mlDulbecco's PBSLee, A, Kessler, J, Read, T, Kaiser, C, Corbeil, D, Huttner ,W, Johnson, J, Wechsler-Reya, R.: Isolation of Neural Stem Cells from the Postnatal Cerebellum, Nat Neurosci 8, 723, 2005
Deoxyribonuclease I: 250 u/ml
小鼠小鼠, 胚胎Embryonic fibroblast feeder cellsCollagenase Type 4: 0.1%DMEMSchatten Gerald, Smith Joseph, Navara Christopher, Park Jong-Hyuk, Pedersen Roger:Culture of human embryonic stem cells, Nat Methods 2, 455-63, 2005
小鼠小鼠, 1天Neural progenitorPDS kit: per instructionssee referenceSeaberg, 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, 8-10 周Liver derived stem cellsCollagenase Type 1: 0.1%HBSSKotton, D., Fabian, A. and Mulligan, R.: A Novel Stem-Cell Population in Adult Liver with Potent Hematopoietic-Reconstitution Activity., Blood 106, 1574, 2005
Neutral Protease: 2.4 u/ml
小鼠小鼠Bone marrowCollagenase Type 1: 0.15%PBSBertoncello, I. and Williams, B.: Hematopoietic Stem Cell Characterization by Hoechst 33342 and Rhodamine 123 Staining., Methods Mol Biol 263, 181, 2004
Neutral Protease: 0.15%
小鼠小鼠, 4-7 天Pluripotent stem cellsCollagenase Type 1: 220 u/mlMEMHowell, J., Lee, W., Morrison, P., Zhong, J., Yoder, M. and Srour, E.: Pluripotent Stem Cells Identified in Multiple Murine Tissues., Ann N Y Acad Sci 996, 158, 2003
Neutral Protease: 33 u/ml
小鼠小鼠, 15 天 或 3 个月Neural stem cellsPapain: 0.1%DMEM/F-12Gritti, A, Bonfanti, L, Doetsch, F, Caille, I, Alvarez-Buylla, A, Lim, D, Galli, R, Verdugo J, Herrera, D, and Vescovi A.: Multipotent Neural Stem Cells Reside into the Rostral Extension and Olfactory Bulb of Adult Rodents, J Neurosci 22(2), 437, 2002
小鼠小鼠, 胚胎Cortical progenitorsPDS kit: per instructionsSerum free mediumEstivill-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, 4-8 个月Neural subventricular zoneTrypsin: 0.13%DMEM/F12Gritti A, Frolichsthal-Schoeller P, Galli R, Parati E, Cova L, Pagano S, Bjornson C, and Vescovi A.: Epidermal and Fibroblast Growth Factors Behave as Mitogenic Regulators for a Single Multipotent Stem Cell-like Population from the Subventricular Region of the Adult Mouse forebrain, J Neurosci 19(9), 3287, 1999
Hyaluronidase: 0.067%
小鼠小鼠, 6 周Muscle hematopoietic stem cellsCollagenase: 0.2%DMEMJackson, K., and Goodell, M.: Hematopoietic Potential of Stem Cells Isolated from Murine Skeletal Muscle, Proc Natl Acad Sci U S A 96, 14482, 1999
Trypsin: 0.1%
猪, 3-4 个月Synovial membrane and skin stem cellsCollagenase: 0.2%DMEMAndo, W., Kutcher, J., Krawetz, R., Sen, A., Nakamura, N., Frank, C. and Hart, D.: Clonal Analysis of Synovial Fluid Stem Cells to Characterize and Identify Stable Mesenchymal Stromal Cell/Mesenchymal Progenitor Cell Phenotypes in a Porcine Model: A Cell Source with Enhanced Commitment to the Chondrogenic Lineage., Cytotherapy 16, 776, 2014
猪,雌性, 小于一岁Adipose mesenchymal stemCollagenase Type 1: 0.1%DMEMWilliams, K., Picou, A., Kish, S., Giraldo, A., Godke, R. and Bondioli, K: Isolation and Characterization of Porcine Adipose Tissue-Derived Adult Stem Cells., Cells Tissues Organs188, 251, 2008
兔, 新西兰, 8-10 周Tenocytes and tendon stem cellsCollagenase Type 1: 0.3%DMEMZhang, J. and Wang, J.: Characterization of Differential Properties of Rabbit Tendon Stem Cells and Tenocytes., BMC Musculoskelet Disord 11, 10, 2010
Neutral Protease: 0.4%
大鼠大鼠Mesenchymal Stem CellsCollagenase Type 1: 0.075%DMEMEmre, E, Yuksel, N, Duruksu, G, Pirhan, D, Subasi, C, Erman, G and Karaoz, E:Neuroprotective Effects of Intravitreally Transplanted Adipose Tissue and Bone Marrow-Derived Mesenchymal Stem Cells in an Experimental Ocular Hypertension Model.,Cytotherapy 17, 543-59, 2015
大鼠SD大鼠, 5 个月Mesenchymal Stromal CellsCollagenase Type 2: 0.075%DMEMVeronesi, F, Torricelli, P, Della, B, Pagani, S and Fini, M: In Vitro Mutual Interaction Between Tenocytes and Adipose-Derived Mesenchymal Stromal Cells., Cytotherapy 17, 215-23, 2015
大鼠大鼠Adipose Derived StemCollagenase Type 1: 0.1%DMEMWei, X., Du, Z., Zhao, L., Feng, D., Wei, G., He, Y., Tan, J., Lee, W., Hampel, H., Dodel, R., Johnstone, B., March, K., Farlow, M. and Du, Y.: IFATS Collection: The Conditioned Media of Adipose Stromal Cells Protect Against Hypoxia-Ischemia-Induced Brain Damage In Neonatal Rats., Stem Cells 27, 478, 2009
大鼠SD大鼠, 新生Neutral stem cells, Schwann cellsTrypsin: 0.25%DMEM/F12Zeng Yuan-Shan, Ding Ying, Wu Li-Zhi, Guo Jia-Song, Li Hai-Biao, Wong Wai-Man, Wu Wu-Tian: Co-transplantation of schwann cells promotes the survival and differentiation of neural stem cells transplanted into the injured spinal cord, Dev Neurosci 27, 20-6, 2005
Collagenase: 0.16%
大鼠大鼠, 雄性, 200-250 克Neural stem cellsPapain: 0.09%EBSSGobbel GT, Choi SJ, Beier S, Niranjan A: Long-term c*tion of multipotential neural stem cells from adult rat subependyma, Brain Res 980, 221, 2003
Deoxyribonuclease I: 0.1%
大鼠大鼠, 出生后的, 成年的Gut Neural Crest Stem CellsCollagenase Type 4: 0.1%HBSSKruger, G., Mosher, J., Bixby, S., Joseph, N., Iwashita, T. and Morrison, S.: Neural Crest Stem Cells Persist in the Adult Gut but Undergo Changes in Self-Renewal, Neuronal Subtype Potential, and Factor Responsiveness., Neuron 35, 657, 2002
Trypsin: 0.025%
大鼠大鼠, 成年的Neural stem cellsPapain: 2.5 u/mlDMEM/F-12Palmer, T., Markakis, E., Willhoite, A., Safar, F., and Gage, F.: Fibroblast Growth Factor-2 Activates a Latent Neurogenic Program in Neural Stem Cells from Diverse Regions of the Adult CNS, J Neurosci 19, 8487, 1999
Deoxyribonuclease I: 250 u/ml
Neutral Protease: 1 u/ml
大鼠大鼠Central nervous system stem cellsPapain: 0.01%DMEM/F-12Ray, J. and Gage, F.: Neural Stem Cell Isolation, Characterization and Transplantation,Modern Techniques in Neuroscience Research,Johansson, H. and Windhorst, U., Springer-Verlag New York, 339, 1999
Neutral Protease: 0.1%
大鼠大鼠, 胚胎Neurons and progenitorPapain: 20 u/mlEBSSMaric, D., Maric, I. and Barker, J.: Buoyant Density Gradient Fractionation and Flow Cytometric Analysis of Embryonic Rat Cortical Neurons and Progenitor Cells., Methods 16, 247, 1998

 

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