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原代細(xì)胞分離中，軟骨組織分離涉及到軟骨細(xì)胞、間充質(zhì)、基質(zhì)細(xì)胞囊泡、關(guān)節(jié)軟骨細(xì)胞、骨骺和關(guān)節(jié)軟骨等等的解離通常會(huì)使用酶，如何選擇合適的酶來(lái)完成對(duì)應(yīng)軟骨組織的解離，以及原代細(xì)胞采集的常規(guī)操作， 艾美捷針對(duì)軟骨組織相關(guān)的組織分類(lèi)解離根據(jù)各種屬、不同的細(xì)胞、酶組合推薦，推薦濃度、培養(yǎng)基以及參考文獻(xiàn)匯總詳細(xì)指南：
本技術(shù)指南描述了標(biāo)準(zhǔn)實(shí)驗(yàn)室程序； 為建立細(xì)胞分離方案提供了一種合乎邏輯的實(shí)驗(yàn)方法； 并列出了許多組織特異性參考資料，以幫助開(kāi)發(fā)有效的方法。更多其他組織的解離指南，點(diǎn)擊查看：各組織分離指南
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| 種屬 | 種屬詳情 | 細(xì)胞名稱(chēng) | 酶組合推薦，含推薦濃度 | 培養(yǎng)基 | 參考文獻(xiàn) |
| 牛 | Bovine, 18-36 month | 軟骨細(xì)胞 / Chondrocytes | Collagenase Type 1: 0.1% | DMEM | White, R. and Gibson, J.: The Effect of Oxygen Tension on Calcium Homeostasis in Bovine Articular Chondrocytes., J Orthop Surg Res 5, 27, 2010 |
| 牛 | Bovine, 6-8 week | 軟骨細(xì)胞 / Chondrocytes | Collagenase Type 2: 0.2% | DMEM | Hwang, Y., Sangaj, N. and Varghese, S.: Interconnected Macroporous Poly(ethylene glycol) Cryogels as a Cell Scaffold for Cartilage Tissue Engineering., Tissue Eng Part A 16, 3033-41, 2010 |
| 牛 | Bovine, 1-3 month | 軟骨細(xì)胞 / Chondrocytes | Collagenase Type 4: 390 u/ml | DMEM | Lima, E., Tan, A., Tai, T., Marra, K., DeFail, A., Ateshian, G. and Hung, C.: Genipin Enhances the Mechanical Properties of Tissue-Engineered Cartilage and Protects Against Inflammatory Degradation When Used as a Medium Supplement., J Biomed Mater Res 91, 692, 2009 |
| 牛 | Steers, 1-2 years | 軟骨細(xì)胞 / Chondrocytes | Trypsin: 0.20% | HBSS | Mackintosh, D., and Mason, R.: Pharmacological Actions of 17 Beta-oestradiol on Articular Cartilage Chondrocytes and Chondrosarcoma Chondrocytes in the Absence of Oestrogen Receptors, Biochim Biophys Acta 964, 295, 1988 |
| 牛 | Calf, 1-14 days | 軟骨細(xì)胞 / Chondrocytes | Collagenase Type 2: 0.20% | PBS | Klagsbrun, M.: Large Scale Preparation of Chondrocytes,Methods of Enzymology Vol. 58,, , 560, 1979 |
| 犬/狗 | Canine | 軟骨細(xì)胞 / Chondrocytes | Collagenase Type 2: 0.3%； Trypsin: 0.25% | Ham's F12 | Lee, J., Kim, H., Kim, J., Bae, S., Joo, D., Huh, K., Fang, Y., Jeong, J., Kim, M. and Kim, Y.: Microencapsulation of Pancreatic Islets with Canine Ear Cartilage for Immunoisolation., Transplant Proc 44, 1091-4, 2012 |
| 雞 | Chick embryos, HH stage | 間充質(zhì) / Mesenchymal | Trypsin: 0.1% | DMEM；(see reference) | Wong, M., and Tuan, R.: Nuserum, A Synthetic Serum Replacement, Supports Chondrogenesis of Embryonic Chick Limb Bud Mesenchymal Cells in Micromass Culture, In Vitro Cell Dev Biol 29A, 917, 1993 |
| 雞 | Chicken, broiler strain, 8-10 weeks | 基質(zhì)細(xì)胞囊泡，骨骺生長(zhǎng)板 / Matrix vesicles，Epiphyseal growth plate | Trypsin: 0.1% | Tris-buffered saline | Genge, B., Wu, L. and Wuthier, R.: Differential Fractionation of Matrix Vesicle Proteins: Further Characterization of the Acidic Phospholipid-dependent Ca2+-Binding Proteins, J Biol Chem 265, 4703, 1990 |
| 雞 | Chicken, broiler strain, 8-10 week | 基質(zhì)細(xì)胞囊泡 / Matrix vesicles | Trypsin: 0.1% | (see reference) | Genge, B.R., Wu, L.N.Y., and Wuthier, R.E.: Identification of Phospholipid-dependent Calcium-binding Proteins as Constituents of Matrix Vesicles, J Biol Chem 264, 10917, 1989 |
| 雞 | Chick embryos, White Leghorn, 19 day old | 成纖維細(xì)胞，上皮樣 / Fibroblasts，Epithelial-like | Trypsin: 0.25% | E 199 medium | Gionti, E., Capasso, O., and Cancedda, R.: The Culture of Chick Embryo Chondrocytes and the Control of Their Differentiated Functions in Vitro, J Biol Chem 258 (11), 7190, 1983 |
| 雞 | Chick embryos, 19 day old | 軟骨細(xì)胞 / Chondrocytes | Trypsin: 0.75% | Coon's modified F-12 | Capasso, O., Gionti, E., Pontarelli, G., Ambesi-Impiobato, F., Nitsch, L., Tajana, G., and Cancedda, R.: The Culture of Chick Embryo Chondrocytes and the Control of Their Differentiated Functions In Vitro, Exp Cell Res 142, 197, 1982 |
| 雞 | Chick embryos, White Leghorn | 翼芽 / Wing buds | Trypsin: 0.1% | Saline G | Ahrens, P., Solursh, M., and Reiter, R.: Stage-Related Capacity for Limb Chondrogenesis in Cell Culture, Dev Biol 60, 69, 1977 |
| 馬 | Equine | 軟骨細(xì)胞 / Chondrocytes | Collagenase Type 2: 0.15% | DMEM | Visser, J., Levett, P., te Moller, N., Besems, J., Boere, K., van Rijen, M., de Grauw, J., Dhert, W., van Weeren, P. and Malda, J.: Crosslinkable Hydrogels Derived from Cartilage, Meniscus, and Tendon Tissue., Tissue Eng Part A 21, 1195-206, 2015 |
| 馬 | Equine | 軟骨細(xì)胞 / Chondrocytes | Collagenase Type 2: 0.08% | DMEM/F12 | Ley, C., Svala, E., Nilton, A., Lindahl, A., Eloranta, M., Ekman, S. and Skioldebrand, E.: Effects of High Mobility Group Box Protein-1, Interleukin-1B, and Interleukin-6 on Cartilage Matrix Metabolism in Three-Dimensional Equine Chondrocyte Cultures., Conn Tissue Res 52, 290-300, 2011 |
| 山羊 | Goat | 軟骨細(xì)胞 / Chondrocytes | Collagenase Type 2: 2% | DMEM | Bekkers,J, Creemers, L, Tsuchida, A, van Rijen, M, Custers, R, Dhert, W and Saris, D.: One-Stage Focal Cartilage Defect Treatment with Bone Marrow Mononuclear Cells and Chondrocytes Leads to Better Macroscopic Cartilage Regeneration Compared to Microfracture in Goats., Osteoarthritis Cartilage 21, 950-6, 2013 |
| 人 | Human | 軟骨細(xì)胞 / Chondrocytes | Trypsin: 0.25%； Hyaluronidase: 0.1%； Collagenase Type 2: 1000 u/ml； Collagenase Type 1: 400 u/ml | Ham's F12 | Muraglia, A., Nguyen, V., Nardini, M., Mogni, M., Coviello, D., Dozin, B., Strada, P., Baldelli, I., Formica, M., Cancedda, R. and Mastrogiacomo, M.: Culture Medium Supplements Derived from Human Platelet and Plasma: Cell Commitment and Proliferation Support., Front Bioeng Biotechnol 5, 66, 2017 |
| 人 | Human | 軟骨細(xì)胞 / Chondrocytes | Collagenase Type 2: 0.15% | DMEM | Andrews, S., Kunze, M., Mulet-Sierra, A., Williams, L., Ansari, K., Osswald, M. and Adesida, A.: Strategies to Mitigate Variability in Engineering Human Nasal Cartilage., Sci Rep 7, 6490, 2017 |
| 人 | Human | 軟骨細(xì)胞 / Chondrocytes | Collagenase Type 2: 0.15% | DMEM | Capsoni, F., Ongari, A., Lonati, C., Accetta, R., Gatti, S. and Catania, A.: Alpha-Melanocyte-Stimulating-Hormone (a-MSH) Modulates Human Chondrocyte Activation Induced by Proinflammatory Cytokines., BMC Musculoskelet Disord 16, 154, 2015 |
| 人 | Human | 軟骨細(xì)胞 / Chondrocytes | Collagenase: 0.3% | DMEM | Tamai, M., Nagasao, T., Yanaga, H., Hamamoto, Y., Kogure, T. and Tanaka, Y.: Correction of Secondary Deformity after Nuss Procedure for Pectus Excavatum by Means of Cultured autologous Cartilage Cell Injection., Int J Surg Case Rep 15, 70-3, 2015 |
| 人 | Human, 52-82 years | 軟骨細(xì)胞 / Chondrocytes | Collagenase Type 2: 0.2%； Pronase: 0.15% | DMEM/F12 | Pallu, S., Francin, P., Guillaume, C., Gegout-Pottie, P., Netter, P., Mainard, D., Terlain, B. and Presle, N.: Obesity Affects the Chondrocyte Responsiveness to Leptin in Patients with Osteoarthritis., Arthritis Res Ther 12, R112, 2010 |
| 人 | Human | 滑膜細(xì)胞 / Synoviocytes | Collagenase Type 1: 0.4% | DMEM | Kim Wan-Uk, Kwok Seung-Ki, Hong Kyung-Hee, Yoo Seung-Ah, Kong Jin-Sun, Choe Jongseon, Cho Chul-Soo: Soluble Fas ligand inhibits angiogenesis in rheumatoid arthritis, Arthritis Res Ther 9, R42, 2007 |
| 人 | Human, 26-68 year | 半月板和軟骨 / Meniscus and cartilage | Collagenase Type 2: 0.15% | DMEM | Marsano A, Millward-Sadler SJ, Salter DM, Adesida A, Hardingham T, Tognana E, Kon E, Chiari-Grisar C, Nehrer S, Jakob M, Martin I: Differential cartilaginous tissue formation by human synovial membrane, fat pad, meniscus cells and articular chondrocytes, Osteoarthritis Cartilage 15, 48-58, 2007 |
| 人 | Human, 64-83 yr | 軟骨細(xì)胞 / Chondrocytes | Collagenase Type 2: 0.08% | DMEM/F12 | Tallheden T, Bengtsson C, Brantsing C, Sjogren-Jansson E, Carlsson L, Peterson L, Brittberg M, and Lindahl A.: Proliferation and Differentiation Potential of Chondrocytes from Osteoarthritic Patients, Arthritis Res Ther 7(3), R560, 2005 |
| 人 | Human, adult | 滑膜 / Synovial | Collagenase: 0.15%； Hyaluronidase: 0.1%； Deoxyribonuclease I: 0.015% | DMEM | Liagre B, Vergne-Salle P, Corbiere C, Charissoux JL, and Beneytout JL.: Diosgenin, a Plant Steroid, Induces Apoptosis in Human Rheumatoid Arthritis Synoviocytes with Cyclooxygenase-2 Overexpression, Arthritis Res Ther 6(4), R373, 2004 |
| 人 | Human, adult | 軟骨細(xì)胞 / Chondrocytes | Collagenase Type 2: 0.15% | DMEM | Jakob M, Demarteau O, Schafer D, Stumm M, Heberer M, and Martin I.: Enzymatic Digestion of Adult Human Articular Cartilage Yields a Small Fraction of the Total Available Cells, Conn Tissue Res 44, 173, 2003 |
| 人 | Human, adults | 滑膜細(xì)胞 / Synoviocytes | Collagenase Type 1: 0.1% | RPMI | McEvoy A., Murphy E., Ponnio T., Conneely O., Bresnihan B., FitzGerald O., and Murphy E.: Activation of Nuclear Orphan Receptor NURR1 Transcription by NF-kappa B and Cyclic Adenosine 5'-Monophosphate Response Element-binding Protein in Rheumatoid Arthritis Synovial Tissue, J Immunol 168(6), 2979, 2002 |
| 人 | Human, 15-60 years | 軟骨細(xì)胞 / Chondrocytes | Collagenase Type 2: 0.2% | DMEM | Rotter N, Bonassar LJ, Tobias G, Lebl M, Roy AK, Vacanti CA: Age dependence of cellular properties of human septal cartilage: implications for tissue engineering, Arch Otolaryngol Head Neck Surg 127, 1248-52, 2001 |
| 人 | Human | 滑膜成纖維細(xì)胞 / Synovial fibroblasts | Collagenase: 0.4% | DMEM | Sarkissian M, Lafyatis R: Integrin engagement regulates proliferation and collagenase expression of rheumatoid synovial fibroblasts, J Immunol 162, 1772-9, 1999 |
| 人 | Human | 隔膜軟骨細(xì)胞 / Septal chondrocytes | Collagenase Type 2: 0.2%； Hyaluronidase: 0.01%； Deoxyribonuclease I: 0.015% | DMEM/F12 | Dunham B P, Koch R J: Basic fibroblast growth factor and insulinlike growth factor I support the growth of human septal chondrocytes in a serum-free environment, Arch Otolaryngol Head Neck Surg 124, 1325-30, 1998 |
| 人 | Human w/ rheumatoid arthritis | 滑膜組織 / Synovial tissue | Trypsin: 0.05% | DMEM (see reference) | Dayer, J., Krane, S., Russell, R., and Robinson, D.: Production of Collagenase and Prostaglandins by Isolated Adherent Rheumatoid Synovial Cells, Proc Natl Acad Sci U S A 73 (3), 945, 1976 |
| 人 | Human, 13-62 years | 關(guān)節(jié)軟骨細(xì)胞 / Articular chondrocytes | Trypsin: 0.2% | BSS | Srivastava, V.M.L., Malemud, C.J., Hough, A.J., Bland, J.H., and Sokoloff, L.: Preliminary Experience with Cell Culture of Human Articular Chondrocytes, Arthritis Rheum 17, 165, 1974 |
| 人 | Human, 26-84 years | 軟骨細(xì)胞 / Chondrocytes | Collagenase: | GBSS | Manning, W.K., and Bonner, W.M.: Isolation and Culture of Chondrocytes From Human Adult Articular Cartilage, Arthritis Rheum 10, 235, 1967 |
| 小鼠 | Mouse, 2-5 day | 軟骨細(xì)胞 / Chondrocytes | Trypsin: 0.25%； Collagenase Type 1: 86.5 u/ml | DMEM | Otsuru, S., Hofmann, T., Raman, P., Olson, T., Guess, A., Dominici, M. and Horwitz, E.: Genomic and Functional Comparison of Mesenchymal Stromal Cells Prepared Using Two Isolation Methods., Cytotherapy 17, 262-70, 2015 |
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