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worthington 皮肤组织分离指南

发布者:艾美捷科技    发布时间:2022-06-09     
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原代细胞分离中,针对皮肤组织涉及到表皮,真皮分离的聚集蛋白聚糖敏感、二倍体成纤维细胞 、皮肤成纤维细胞,皮肤和角质形成细胞等等的解离通常会使用酶,如何选择合适的酶来完成对应皮肤组织的解离,以及原代细胞采集的常规操作, 艾美捷针对皮肤相关的组织分类解离分种属、不同的细胞、酶组合推荐,推荐浓度、培养基以及参考文献,详细汇总指南:皮肤组织分离指南.png

 

本技术指南描述了标准实验室程序; 为建立细胞分离方案提供了一种合乎逻辑的实验方法; 并列出了许多组织特异性参考资料,以帮助开发有效的方法。更多其他组织的解离指南,点击查看:各组织分离指南

 

皮肤
种属种属详情细胞名称酶组合推荐,含推荐浓度培养基参考文献
犬/狗Canine表皮 / EpidermalCollagenase: 0.35%;
Neutral Protease: 0.1%
DMEMZheng, Y., Nace, A., Chen, W., Watkins, K., Sergott, L., Homan, Y., Vandeberg, J., Breen, M. and Stenn, K.: Mature Hair Follicles Generated from Dissociated Cells: A Universal Mechanism of Folliculoneogenesis., Dev Dyn 239, 2619-26, 2010
青蛙Frog表皮 / EpidermalTrypsin: 0.18%Barth's solution, CMFNishikawa, A., Shimizu-Nishikawa, K., and Miler, L.: Isolation, characterization, and in vitro culture of larval and adult epidermal cells of the frog Xenopus laevis, In Vitro Cell Dev Biol 26, 1128, 1990
山羊Goat真皮分离的聚集蛋白聚糖敏感 / Dermis isolated aggrecan sensitiveNeutral Protease: 0.5%;
Collagenase Type 2: 200 u/ml
DMEMDeng, Y., Hu, J. and Athanasiou, K.: Isolation and Chondroinduction of a Dermis-Isolated, Aggrecan-Sensitive Subpopulation with High Chondrogenic Potential., Arthritis Rheum 56, 168, 2007
Human二倍体成纤维细胞 / Diploid fibroblastsCollagenase Type 1: 0.03%DMEMTan, J., Jaafar, F. and Makpol, S.: Proteomic Profiling of Senescent Human Diploid Fibroblasts Treated with gamma-tocotrienol., BMC Complement Altern Med 18, 314, 2018
Human成纤维细胞 / FibroblastsNeutral Protease: 0.1%;
Collagenase Type 1: 0.25%
DMEMChen, Z., Wang, X., Jin, T., Wang, Y., Hong, C., Tan, L., Dai, T., Wu, L., Zhuang, Z. and Shi, C.: Increase in the Radioresistance of Normal Skin Fibroblasts but not Tumor Cells by Mechanical Injury., Cell Death Dis 8, e2573, 2017
Human单核吞噬细胞 / Mononuclear phagocytes

Neutral Protease: 1 u/ml;

Deoxyribonuclease I: 50 u/ml;

Collagenase: 200 u/ml;

Trypsin: 0.5%;

RPMI-1640Botting, Ra., Bertram, K., Baharlou, H., Sandgren, K., Fletcher, J., Rhodes, J., Rana, H., Plasto, T., Wang, X., Lim, J., Barnouti, L., Kohout, M., Papadopoulos, T., Merten, S., Olbourne, N. and Cunningham, A.: Phenotypic and Functional Consequences of Different Isolation Protocols on Skin Mononuclear Phagocytes., J Leukoc Biol 101, 1393-1403, 2017
Human皮肤 / Skin

Neutral Protease: 5 u/ml;

Collagenase Type 3: 0.3%;

Deoxyribonuclease I: 0.005%;

RPMI 1640Cheuk, S., Wiken, M., Blomqvist, L., Nylen, S., Talme, T., Stahle, M.and Eidsmo, L.: Epidermal Th22 and Tc17 Cells Form a Localized Disease Memory in Clinically Healed Psoriasis., J Immunol 192, 3111, 2014
Human皮肤成纤维细胞 / Skin fibroblastsCollagenase Type 1: 1000 u/mlDMEMDouvaras, P., Wang, J., Zimmer, M., Hanchuk, S., O'Bara, M., Sadiq, S., Sim, F., Goldman, J. and Fossati, V.: Efficient Generation of Myelinating Oligodendrocytes from Primary Progressive Multiple Sclerosis Patients by Induced Pluripotent Stem Cells., Stem Cell Reports 3, 250, 2014
Human皮肤 / Skin

Collagenase Type 4: 0.08-0.2%;

Neutral Protease: 0.1%;

Deoxyribonuclease I: 0.005%;

RPMICerny, D., Haniffa, M., Shin, A., Bigliardi, P., Tan, B., Lee, B., Poidinger, M., Tan, E., Ginhoux, F. and Fink, K.: Selective Susceptibility of Human Skin Antigen Presenting Cells to Productive Dengue Virus Infection., PLoS Pathog 10, e1004548, 2014
Human, adult表皮干细胞 / Epidermal stem cellsNeutral Protease: 2 u/ml;
Trypsin: 2.5%
DMEMNagel, 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
Human成纤维细胞 / FibroblastsCollagenase animal free: 2%HBSSKarumbayaram, S., Lee, P., Azghadi, S., Cooper, A., Patterson, M., Kohn, D., Pyle, A., Clark, A., Byrne, J., Zack, J., Plath, K. and Lowry, W.: From Skin Biopsy to Neurons Through a Pluripotent Intermediate Under Good Manufacturing Practice Protocols, Stem Cells Transl Med 1, 36, 2012
Human, female, 18-66 yr成纤维细胞 / FibroblastsTrypsin: 0.2%DMEMHuschtscha, L., Napier, C., Noble, J., Bower, K., Au, A., Campbell, H., Braithwaite, A. and Reddel, R.: Enhanced Isolation of Fibroblasts From Human Skin Explants., Biotechniques 53, 239, 2012
Human成纤维细胞和角质形成细胞 / Fibroblasts and keratinocytesNeutral Protease: 0.5%;
Collagenase Type 1: 0.2%
DMEMCarriel, V., Garzon, I., Jimenez, J., Oliveira, A., Arias-Santiago, S., Campos, A., Sanchez-Quevedo, M. and Alaminos, M.: Epithelial and Stromal Developmental Patterns in a Novel Substitute of the Human Skin Generated with Fibrin-Agarose Biomaterials., Cells Tissues Organs 196, 1-12, 2012
Human表皮 / EpidermalCollagenase: 0.35%;
Neutral Protease: 0.1%
DMEMZheng, Y., Nace, A., Chen, W., Watkins, K., Sergott, L., Homan, Y., Vandeberg, J., Breen, M. and Stenn, K.: Mature Hair Follicles Generated from Dissociated Cells: A Universal Mechanism of Folliculoneogenesis., Dev Dyn 239, 2619-26, 2010
Human皮肤和瘢痕疙瘩祖细胞 / Skin and keloid progenitorCollagenase Type 1: 0.4%;
Neutral Protease: 0.3%
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
Human成纤维细胞 / FibroblastsCollagenase Type 1: 100-320 u/mlDMEMTuan, T., Hwu, P., Ho, W., Yiu, P., Chang, R., Wysocki, A. and Benya, P.: Adenoviral Overexpression and Small Interfering RNA Suppression Demonstrate that Plasminogen Activator Inhibitor-1 Produces Elevated Collagen Accumulation in Normal and Keloid Fibroblasts., Am J Pathol 173, 1311-25, 2008
Human, 6-12 year真皮成纤维细胞 / Dermal fibroblastsNeutral Protease: 0.1%;
Collagenase Type 1: 0.1%
DMEMChen, F., Zhang, W., Bi, D., Liu, W., Wei, X., Chen, F., Zhu, L., Cui, L. and Cao, Y.: Clonal Analysis of Nestin(-) Vimentin(+) Multipotent Fibroblasts Isolated from Human Dermis., J Cell Sci 120, 2875, 2007
Human毛囊上皮 / Hair follicular epithelialNeutral Protease: 1.25%;
Trypsin: 0.25%
DMEM/F-12Yu Hong, Fang Dong, Kumar Suresh M, Li Ling, Nguyen Thiennga K, Acs Geza, Herlyn Meenhard, Xu Xiaowei: Isolation of a novel population of multipotent adult stem cells from human hair follicles, Am J Pathol 168, 1879-88, 2006
Human成纤维细胞 / fibroblastsCollagenase: 0.5%;
Trypsin: 0.25%
DMEM/F12Clark, R., Chong, B., Mirchandani, N., Yamanaka, K., Murphy, G., Dowgiert, R. and Kupper, T.: A Novel Method for the Isolation of Skin Resident T cells from Normal and Diseased Human Skin., J Invest Dermatol 126, 1059, 2006
Human, neonatal角质形成细胞 / KeratinocytesNeutral Protease: 0.4%;
Collagenase: 0.3%
DMEMLi, A., Pouliot, N., Redvers, R., and Kaur, P.: Extensive tissue-regenerative capacity of neonatal human keratinocytes stem cells and their progeny, J Clin Invest 113, 390-400, 2004
Human肥大细胞 / Mast cellsNeutral Protease: 0.05%;
Collagenase Type 4: 1.0%
RPMIBabina Magda, Guhl Sven, Starke Andrae, Kirchhof Loreen, Zuberbier Torsten, Henz Beate M: Comparative cytokine profile of human skin mast cells from two compartments--strong resemblance with monocytes at baseline but induction of IL-5 by IL-4 priming, J Leukoc Biol 75, 244-52, 2004
Human, adult真皮成纤维细胞 / Dermal fibroblastsCollagenase: 0.5-1.0%DMEMWang, H., Van Blitterswijk, C., Bertrand-De Haas, M., Schuurman, A. and Lamme, E.: Improved Enzymatic Isolation of Fibroblasts for the Creation of Autologous Skin Substitutes., In Vitro Cell Dev Biol Anim 40, 268, 2004
Human成纤维细胞 / FibroblastsTrypsin: 0.25%DMEMTuan Tai-Lan, Wu Huayang, Huang Eunice Y, Chong Sheree S N, Laug Walter, Messadi Diana, Kelly Paul, Le Anh: Increased plasminogen activator inhibitor-1 in keloid fibroblasts may account for their elevated collagen accumulation in fibrin gel cultures, Am J Pathol 162, 1579-89, 2003
Human角质形成细胞,成纤维细胞,内皮细胞 / Keratinocytes, fibroblasts, endothelialNeutral Protease: see reference;
Collagenase: see reference
see referenceSupp Dorothy M, Wilson-Landy Kaila, Boyce Steven T: Human dermal microvascular endothelial cells form vascular analogs in cultured skin substitutes after grafting to athymic mice, FASEB J 16, 797-804, 2002
Human, adult角质形成细胞 / KeratinocytesNeutral Protease: 0.25%PBSBaudoux, B., Castanares-Zapatero, D., Leclercq-Smekens, M., Berna, N., and Poumay, Y.: The Tetraspanin CD9 Associates with the Integrin a6B4 in Cultured Human Epidermal Keratinocytes and is Involved in Cell Motility, Eur J Cell Biol 79, 41, 2000
Human角质形成细胞 / Keratinocytes

Neutral Protease: 0.25%;

Trypsin: 0.17%;

Thermolysin: 0.05%;

see referenceHybbinette S, Bostrom M, Lindberg K: Enzymatic dissociation of keratinocytes from human skin biopsies for in vitro cell propagation, Exp Dermatol 8, 30-8, 1999
Human人皮肤肥大细胞 / Human skin mast cellsNeutral Protease: 0.1%;
Collagenase Type 1: see reference
PBSGrutzkau A, Kruger-Krasagakes S, Baumeister H, Schwarz C, Kogel H, Welker P, Lippert U, Henz BM, Moller A: Synthesis, storage, and release of vascular endothelial growth factor/vascular permeability factor (VEGF/VPF) by human mast cells: implications for the biological significance of VEGF206, Mol Biol Cell 9, 875-84, 1998


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