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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(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*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。
干細(xì)胞組織消化手冊
| 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 | 人 | Adipose derived stem cells | Collagenase Type 1: 0.1% | DMEM | Tandon, 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 |
| 犬 | 犬 | Amniotic membrane MSC | Trypsin: 0.25% | LG-DMEM | Park, 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 | 犬, 20-25 公斤 | Adipose stem cells | Collagenase: see reference | Media-199 | Fischer, 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 |
| 馬 | 馬 | Adipose derived stem cells | Collagenase Type 1: 0.1% | PBS | Vidal, 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 |
| 馬 | 馬, 1-5 歲 | Adipose derived stem cells | Collagenase Type 1: 0.1% | PBS | Vidal, 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 |
| 人類 | 人類 | Mesenchymal Stromal Cells | Collagenase Type 4: 0.2% | PBS | Choudhery,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 |
| 人類 | 人類 | Adipose derived stem cells | Collagenase Type 2: 0.1% | HBSS | Satish, 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 |
| 人類 | 人類 | Dental mesenchymal stem | Collagenase Type 1: 0.3% | DMEM | Wu, 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% | |||||
| 人類 | 人類 | Decidua Mesenchymal Stem | Trypsin: 0.25% | PBS | Kusuma, 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% | |||||
| 人類 | 人類, 成年人 | Adipose derived mesenchymal stem | Collagenase Type 2: 0.1% | see reference | Al-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 |
| 人類 | 人類, 成年人 | Adipose derived stem cells | Collagenase Type 1: 0.15% | DMEM | Koellensperger, 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 |
| 人類 | 人類 | Umbilical mesenchymal stem | Collagenase Type 2: 0.1% | DMEM | Hang, 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偽 Transduction.,PLoS ONE 9, e104133, 2014 |
| Trypsin: 0.25% | |||||
| 人類 | 人類 | Cardiac stem | Collagenase 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 |
| 人類 | 人類 | Adipose stromal stem | Collagenase animal free: 200 u/ml | DMEM/Hams F-12 | Carvalho, 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 |
| 人類 | 人類, 成年人 | Epidermal stem cells | Neutral Protease: 2 u/ml | DMEM | Nagel, 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% | |||||
| 人類 | 人類 | Dental pulp stem | Collagenase Type 1: 0.3% | DMEM/F12 | Bonnamain, 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% | |||||
| 人類 | 人類 | Umbilical Cord Mesenchymal Stem | Collagenase: 0.1-0.2% | DMEM | Han, 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% | |||||
| 人類 | 人類 | Dental pulp derived stem cell | Collagenase Type 1: 0.3% | DMEM | Sakai, 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% | |||||
| 人類 | 人類 | Adipose-derived stem cells | Collagenase Type 1: 0.1% | PBS | Cervelli, 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 |
| 人類 | 人類 | Mesenchymal stem | Collagenase Type 1: 0.4% | DMEM/F12 | Shalini, 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% | |||||
| 人類 | 人類 | Wharton jelly mesenchymal stem | Collagenase Type 2: 0.1% | DMEM | Salehinejad, 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% | |||||
| 人類 | 人類 | Placental Mesenchymal Stem | Collagenase Type 2: 0.4% | DMEM | Vellasamy, 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% | |||||
| 人類 | 人類, 男性 40-60 years | Adipose derived stem cells | Collagenase: 0.25% | PBS | Blasi, 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% | |||||
| 人類 | 人類 | Umbilical cord stromal stem | Collagenase Type 4: 0.08% | DMEM | Farias, 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% | |||||
| 人類 | 人類 | Vascular wall-resident multipotent stem cells | Collagenase Type 2: 0.2% | see reference | Klein, 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 | |||||
| 人類 | 人類 | Adipose derived stem | Collagenase Type 1: 0.1% | DMEM/Ham's F-12 | Yu, 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 |
| 人類 | 人類 | Glioma stem cells | PDS kit: per instructions | neurobasal medium | Hjelmeland, 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 |
| 人類 | 人類 | Dental pulp and apical papilla stem cells | Collagenase Type 1: 0.3% | MEM | Huang, 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% | |||||
| 人類 | 人類 | Corneal stromal stem | Neutral Protease: 1.2 u/ml | DMEM | Du, 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% | |||||
| 人類 | 人類 | Adipose tissue-derived stem | Collagenase Type 2: 1.0% | DMEM/F12 | Tan, 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 |
| 人類 | 人類 | Umbilical cord mesenchymal | Hyaluronidase: 0.05% | DMEM | Zeddou, 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% | |||||
| 人類 | 人類 | Adipose stem cells | Collagenase Type 1: 0.1% | DMEM | Hareendran, 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 |
| 人類 | 人類, 40-65 year | Adult human adipose stem cells | Collagenase Type 2: 0.075% | DMEM | Sun, 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 |
| 人類 | 人類 | Filum terminale neural progenitor | Trypsin: see reference | DMEM/F-12 | Varghese, 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 |
| 人類 | 人類 | skin and keloid progenitor cells | Collagenase Type 1: 0.4% | PBS | Zhang, 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% | |||||
| 人類 | 人類, 男性 | Spermatogonial stem cells | Collagenase: 1% | DMEM | Kossack, 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% | |||||
| 人類 | 人類 | Muscle derived multiprogenitor cells | Collagenase Type 2: 0.05% | DMEM | Nesti, 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 |
| 人類 | 人類 | Adipose derived stem cells | Collagenase Type 1: 0.075% | DMEM | Jeong, J.: Adipose Stem Cells as a Clinically Available and Effective Source of Adult Stem Cell Therapy, Int J Stem Cells 1, 43, 2008 |
| 人類 | 人類 | Hepatic stem cells and heptoblasts | Collagenase Type 4: 0.014-0.06% | various | Wauthier, 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 |
| 人類 | 人類 | Umbilical mesenchymal stem | Collagenase: 0.1% | DMEM | Secco, 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 |
| 人類 | 人類 | Stem Cells | Collagenase: 280 u/ml | D-PBS | Pilgaard, 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 |
| 人類 | 人類 | Bone marrow derived MSC | Trypsin: 0.05% | DMEM | Welter, 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% | |||||
| 人類 | 人類, 8-12 year | Tendon stem/progenitor | Collagenase Type 1: 0.3% | DMEM | Bi 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% | |||||
| 人類 | 人類, 男性 | Adipose derived adult stem cells | Collagenase Type 1: 0.1% | DMEM | Lei, 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, 成年人 | Adipose-derived adult stem | Collagenase Type 1: 0.1% | DMEM/F-12 Ham's | Mitchell 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 |
| 人類 | 人類 | Mesenchymal stem | Collagenase Type 1: 0.075% | DMEM | Kern 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 |
| 人類 | 人類 | Placental mesenchymal stem | Collagenase Type 2: 270 u/ml | MEM | Portmann-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 | |||||
| 人類 | 人類 | Stromal stem cells | Collagenase Type 1: 0.2% | HBSS | Boquest 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 |
| 人類 | 人類, 女性 | Adipose derived adult stem cells | Collagenase Type 1: 0.1% | DMEM/F-12 | Guilak, 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 |
| 人類 | 人類 | Adult stem cells | Collagenase Type 1: 0.1% | PBS | Devireddy, 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 |
| 人類 | 人類, 女性 | Adipose derived adult stem cells | Collagenase Type 1: 0.1% | DMEM-Ham's F-12 | Aust, 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 |
| 人類 | 人類, 成年人, 男性 | Human epidermal keratinocyte stem cells | Neutral Protease: 0.5% | DMEM | Papini 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 |
| 人類 | 人類 | Umbilical vein mesenchymal stem cells | Collagenase: 1% | PBS | Covas 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 |
| 人類 | 人類, 7-8 歲 | Stem cells Human exfoliated deciduous teeth | Collagenase Type 1: 0.3% | see reference | Miura, 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% | |||||
| 人類 | 人類, 胎兒 | Epithelial progenitor | Collagenase: 0.03% | DMEM | Malhi, 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 |
| 人類 | 人類, 成年人 | Human skin mast cells | Collagenase Type 2: 0.15% | HBSS | Kambe, 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% | |||||
| 人類 | 人類 | Central nervous system stem | Collagenase: 0.1% | HBSS | Uchida, 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% | |||||
| 人類 | 人類 | Embryonic stem | Neutral Protease: 1% | DMEM | Thomson 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% | |||||
| 人類 | 人類 | Muscle-derived stem cells | Trypsin: 0.25% | DMEM/F12 | Alessandri 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, |
| 猴 | 猴 | Embryonic stem | Collagenase Type 4: 0.08% | DMEM | Chen, 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 |
| 猴 | Baboons, 1歲, 7歲, 14 歲 | Primate spermatogonial | Collagenase Type 2: 0.1% | DMEM | Nagano, 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 progenitor | Collagenase Type 1: 0.1% | DMEM | Wang, 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, P4 | Enteric neural crest progenitors | Neutral Protease: 0.5% | DMEM/F12 | Hotta, 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 Stem | Trypsin: 0.25% | HBSS | Jensen, 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 mesenchymal | Collagenase: 0.1% | DMEM | Futami, 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 cells | Papain: 0.01% | DMEM/F12 | Bracko, 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 stem | Collagenase Type 1: 0.025% | HBSS | Sugii, 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 endothelial | Collagenase Type 4: 0.3-1.0% | PBS | Shi, 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 stem | Collagenase Type 2: 0.2% | HBSS | Chow, 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 Cells | Collagenase Type 4: 0.2% | PBS | Janebodin, 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 progenitor | Collagenase Type 2: 0.2% | HBSS | Han, 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 stem | Collagenase Type 1: 0.25% | RPMI 1640 | Xu, 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 stem | Trypsin: 0.05% | DMEM | Deleyrolle, 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/stem | Collagenase Type 2: 0.5% | HBSS | Burger, 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 stem | Collagenase Type 1: 0.075% | Modified Eagles | Cho, 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 cells | Collagenase Type 1: 100 u/ml | HBSS | Breault, 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 cell | Papain: 10 ul/ml | PBS | Hutton, 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 stem | Collagenase Type 1: 0.2% | DMEM/F12 | Staszkiewicz, 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 cells | Trypsin: 0.05% | DMEM | Lu 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/progenitor | Collagenase Type 1: 0.3% | DMEM | Bi 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 cells | Collagenase Type 4: 0.1% | DMEM | Li 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 stem | Collagenase: 0.2% | PBS | Di 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 cells | Papain: see reference | DMEM/F-12 | Meletis, 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 cells | Papain: 10u/ml | Dulbecco's PBS | Lee, 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 cells | Collagenase Type 4: 0.1% | DMEM | Schatten 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 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, 8-10 周 | Liver derived stem cells | Collagenase Type 1: 0.1% | HBSS | Kotton, 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 marrow | Collagenase Type 1: 0.15% | PBS | Bertoncello, 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 cells | Collagenase Type 1: 220 u/ml | MEM | Howell, 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 個月 | Neural stem cells | Papain: 0.1% | DMEM/F-12 | Gritti, 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 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, 4-8 個月 | Neural subventricular zone | Trypsin: 0.13% | DMEM/F12 | Gritti 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 cells | Collagenase: 0.2% | DMEM | Jackson, 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% | |||||
| 豬 | 豬, 3-4 個月 | Synovial membrane and skin stem cells | Collagenase: 0.2% | DMEM | Ando, 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 |
| 豬 | 豬,雌性, 小于一歲 | Adipose mesenchymal stem | Collagenase Type 1: 0.1% | DMEM | Williams, 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 | 兔, 新西蘭, 8-10 周 | Tenocytes and tendon stem cells | Collagenase Type 1: 0.3% | DMEM | Zhang, 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 Cells | Collagenase Type 1: 0.075% | DMEM | Emre, 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 個月 | Mesenchymal Stromal Cells | Collagenase Type 2: 0.075% | DMEM | Veronesi, 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 Stem | Collagenase Type 1: 0.1% | DMEM | Wei, 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 cells | Trypsin: 0.25% | DMEM/F12 | Zeng 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 cells | Papain: 0.09% | EBSS | Gobbel 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 Cells | Collagenase Type 4: 0.1% | HBSS | Kruger, 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 cells | Papain: 2.5 u/ml | DMEM/F-12 | Palmer, 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 cells | Papain: 0.01% | DMEM/F-12 | Ray, 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 progenitor | Papain: 20 u/ml | EBSS | Maric, 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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