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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和細胞生物學研究的原代細胞的分離和收集。盡管這些消化酶已經普遍應用多年，但是對其組織消化和細胞收集的作用機制卻沒有全面的理解。 由此導致的結果便是選擇一種消化方法大多是隨機選擇或基于過去的經驗，而不是在*理解消化原理的基礎上選擇的消化方案，以及如何根據具體實驗目的修正消化方案以獲得更好的實驗結果。
細胞分離程序的目的是zui大程度的獲得功能活性的單個細胞，而影響結果的原因有很多，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、所用的消化培養液
6、所用的酶
7、消化酶中的雜質
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驗決定以獲取zui適實驗結果。
通過查閱文獻來選擇*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據搞得一頭霧水。這些巨大差異來源于組織胞外基質的復雜而動態的特性，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。
本指南總結了目前常用組織消化酶的zui適消化和細胞收集知識和方法，描述了標準的實驗步驟， 以便于制定一個*的細胞分離方案，同時列舉了一些相關的參考文獻。
下表列出多種組織的適宜消化酶（含參考文獻，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 | 結腸組織 | 內皮組織 | 上皮組織 |
眼 | 心臟 | 腸 | 腎 |
肝 | 肺 | 淋巴結 | 乳腺 |
其它組織 | 其他組織 | 肌組織 | 神經組織 |
胰腺 | 腮腺 | 垂體 | 前列腺 |
生殖 | 動物鱗片 | 皮膚 | 脾臟 |
干細胞 | 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。
皮膚組織消化手冊
| 物種 | 詳細來源 | 細胞名稱 | 消化用酶 | 培養基 | 參考文獻 |
| 犬 | 犬 | Epidermal | Collagenase: 0.35% | DMEM | Zheng, 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 |
| Neutral Protease: 0.1% | |||||
| 青蛙 | 青蛙 | Epidermal | Trypsin: 0.18% | Barth's solution, CMF | Nishikawa, 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 | 山羊 | Dermis isolated aggrecan sensitive | Neutral Protease: 0.5% | DMEM | Deng, 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 |
| Collagenase Type 2: 200 u/ml | |||||
| 人類 | 人類 | Skin | Neutral Protease: 5 u/ml | RPMI 1640 | Cheuk, 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 |
| Collagenase Type 3: 0.3% | |||||
| Deoxyribonuclease I: 0.005% | |||||
| 人類 | 人類 | Skin Fibroblasts | Collagenase Type 1: 1000 u/ml | DMEM | Douvaras, 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 Reports3, 250, 2014 |
| 人類 | 人類, 成年的 | 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 | Fibroblasts | Collagenase animal free: 2% | HBSS | Karumbayaram, 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 |
| 人類 | 人類, 女性, 18-66 歲 | Fibroblasts | Trypsin: 0.2% | DMEM | Huschtscha, 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 |
| 人類 | 人類 | Epidermal | Collagenase: 0.35% | DMEM | Zheng, 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 |
| Neutral Protease: 0.1% | |||||
| 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 | Fibroblasts | Collagenase Type 1: 100-320 u/ml | DMEM | Tuan, 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 |
| 人類 | 人類, 6-12 歲 | Dermal fibroblasts | Neutral Protease: 0.1% | DMEM | Chen, 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 Sci120, 2875, 2007 |
| Collagenase Type 1: 0.1% | |||||
| 人類 | 人類 | Hair follicular epithelial | Neutral Protease: 1.25% | DMEM/F-12 | Yu 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 |
| Trypsin: 0.25% | |||||
| 人類 | 人類 | fibroblasts | Collagenase: 0.5% | DMEM/F12 | Clark, 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 |
| Trypsin: 0.25% | |||||
| 人類 | 人類, 新生的 | Keratinocytes | Neutral Protease: 0.4% | DMEM | Li, 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 |
| Collagenase: 0.3% | |||||
| 人類 | 人類 | Mast cells | Neutral Protease: 0.05% | RPMI | Babina 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 |
| Collagenase Type 4: 1.0% | |||||
| 人類 | 人類, 成年的 | Dermal fibroblasts | Collagenase: 0.5-1.0% | DMEM | Wang, 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 |
| 人類 | 人類 | Fibroblasts | Trypsin: 0.25% | DMEM | Tuan 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 |
| 人類 | 人類 | Keratinocytes, fibroblasts, endothelial | Neutral Protease: see reference | see reference | Supp 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 |
| Collagenase: see reference | |||||
| 人類 | 人類, 成年的 | Keratinocytes | Neutral Protease: 0.25% | PBS | Baudoux, 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 |
| 人類 | 人類 | Keratinocytes | Neutral Protease: 0.25% | see reference | Hybbinette S, Bostrom M, Lindberg K: Enzymatic dissociation of keratinocytes from human skin biopsies for in vitro cell propagation, Exp Dermatol 8, 30-8, 1999 |
| Trypsin: 0.17% | |||||
| Thermolysin: 0.05% | |||||
| 人類 | 人類 | Human skin mast cells | Neutral Protease: 0.1% | PBS | Grutzkau 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 |
| Collagenase Type 1: see reference | |||||
| 人類 | 人類 | Epidermal | Collagenase Type 2: 0.1% | PBS | Reece J C, Handley A J, Anstee E J, Morrison W A, Crowe S M, Cameron P U: HIV-1 selection by epidermal dendritic cells during transmission across human skin, J Exp Med 187, 1623-31, 1998 |
| Neutral Protease: 0.5-1.0% | |||||
| 人類 | 人類 | Keratinocytes | Trypsin: 0.05% | Dulbecco's PBS | Judd, D., Battista, P., and Behm, D.: Culture of Human Keratinocytes in Defined Serum-Free Medium, Focus 19 (1), 2, 1997 |
| 人類 | 人類 | Epidermis plus dermis | Trypsin: 0.125% | DMEM, M199 | Harley, C., and Sherwood, S.: Methods in Molecular Biology, Basic Cell Culture Protocols, 2nd ed. Vol. 75,Pollard, J., and Walker, J., Humana Press, 23, 1997 |
| Abdomen or inner forearm | |||||
| 人類 | 人類 | Keratinocytes | Trypsin: 0.25% | DMEM | Regnier, M: Culture of Human Karatinocytes,Cell & Tissue Culture: Laboratory Procedures Vol. 1,Doyle, A., Griffiths, J., and Newell, D., John Wiley and Sons Ltd, 11B:4.1, 1995 |
| 人類 | 人類, 成年的, 女性, 乳房皮膚 | Keratinocytes | Trypsin: 0.25% | PBS | Hirel, B., Chesne, C., Pailgheret, J., and Guillouzo, A.: Expression of Differentiation Markers in Human Adult Keratinocytes Cultured in Submerged Conditions, In Vitro Cell Dev Biol 30A, 372, 1994 |
| 人類 | 人類, 新生兒包皮 | Fibroblasts | Trypsin: 0.25% | DMEM | Hansbrough, J., Cooper, M., Cohen, R., Spielvogel, R., Greenleaf, G., Bar, R., and Naughton, G.: Evaluation of a Biodegradable Matrix Containing Cultured Human Fibroblasts as a Dermal Replacement Beneath Meshed Skin Grafts on Athymic Mice, Surgery 111 (1), 438, 1992 |
| 人類 | 人類, 18-50 歲 | Sweat duct | Collagenase: 0.03% | MEM | Bell, C. and Quinton, P.: Effects of Media Buffer Systems on Growth and Electrophysiologic Characteristics of Cultured Sweat Duct Cells, In Vitro Cell Dev Biol 27, 47, 1991 |
| 人類 | 人類 | Fibroblasts | Trypsin: 0.25% | CMF solution | Limat, A., Hunziker, T., Boillat, C., Noser, F., and Wiesmann, U.: Postmitotic Human Dermal Fibroblasts Preserve Intact Feeder Properties for Epithelial Cell Growth After Long-Term Cryopreservation, In Vitro Cell Dev Biol 26, 709, 1990 |
| 人類 | 人類 | Sweat gland | Collagenase Type 2: 0.015% | (see reference) | Krouse, M., Hagiqara, G., Chen, J., Lewiston, N., and Wine, J.: Ion Channels in Normal Human and Cystic Fibrosis Sweat Gland Cells, Am J Physiol 257, C129, 1989 |
| 人類 | 人類, 年齡16-30 | Human sweat duct | Collagenase Type 2: 0.2% | RPMI 1640 | Pedersen, P.: Human Sweat Duct Cells in Primary Culture. Basic Bioelectric Properties of Cultures Derived From Normals and Patients with Cystic Fibrosis, In Vitro Cell Dev Biol 25 (4), 342, 1989 |
| (see reference) | |||||
| 人類 | 人類 | Melanocytes, skin/ foreskin | Trypsin: 0.25% | PBS | Peacocke, M., Yaar, M., Mansur, C., Chao, M., and Gilchrest, B.: Induction of Nerve Growth Factor Receptors on Cultured Human Melanocytes, Proc Natl Acad Sci U S A 85, 5282, 1988 |
| 人類 | 人類 | Human sweat glands | Collagenase: 0.2% | HBSS | Lee, C., Carpenter, F., Coaker, T., and Kealey, T.: The Primary Culture of Epithelia From the Secretory Coil and Collecting Duct of Normal Human and Cystic Fibrotic Eccrine Sweat Glands, J Cell Sci 83, 103, 1986 |
| 人類 | 人類 | Keratinocytes | Trypsin: 0.25% | DMEM | Dover, R. and Potten, C.: Cell Cycle Kinetics of Cultured Human Epidermal Keratinocytes, J Invest Dermatol 80, 423, 1983 |
| 人類 | 人類, 成年的 | Epidermal keratinocytes | Trypsin: 0.25% | Eagle's MEM | Alitalo K, Kuismanen E, Myllyla R, Kiistala U, Asko-Seljavaara S, Vaheri A: Extracellular matrix proteins of human epidermal keratinocytes and feeder 3T3 cells, J Cell Biol 94, 497-505, 1982 |
| Collagenase: 0.2% | |||||
| Deoxyribonuclease I: 0.001% | |||||
| 人類 | 人類, fetal | Smooth muscle, fibroblasts | Trypsin: 0.055% | DMEM | Davies, P. and Kerr, C.: Modification of LDL Metabolism by Growth Factors in Cultured Vascular Cells and Human Skin Fibroblasts, Biochim Biophys Acta 712, 26, 1982 |
| 人類 | 人類, 18-30歲, 男性, 女性 | Fibroblasts | Trypsin: 0.1% | HBSS | McCoy, B., Galdun, J., and Cohen, I.: Effects of Density and Cellular Aging On Collagen Synthesis and Growth Kinetics in Keloid and Normal Skin Fibroblasts, In Vitro 18 (1), 79, 1982 |
| 人類 | 人類, 新生兒 | Keratinocytes | Trypsin: 0.3% | DMEM | Liu, S., Eaton, M., and Karasek, M.: Growth Characteristics of Human Epidermal Keratinocytes from Newborn Foreskin in Primary and Serial Cultures, In Vitro 15 (10), 813, 1979 |
| 人類 | 人類 | Keratinocytes | Trypsin: 0.25% | (see reference) | Rheinwald, J., and Green, H.: Serial C*tion of Strains of Human Epidermal Kerotinocytes: The Formation of Keratinizing Colonies from Single Cells, Cell 6, 331, 1975 |
| 小鼠 | 小鼠 | Epidermal | Neutral Protease: 0.5% | DMEM | Gu, D., Fan, Q and Xie, J.: Cell Population Analyses During Skin Carcinogenesis., J Vis Exp , e50311, 2013 |
| Collagenase Type 4: 0.1% | |||||
| 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 | Dermal fibroblasts | Collagenase: 400 u/ml | AminoMAX II | Scheibye-Knudsen, M., Ramamoorthy, M., Sykora, P., Maynard, S., Lin, P., Minor, R., Wilson, D., Cooper, M., Spencer, R., de Cabo, R., Croteau, D. and Bohr, V.: Cockayne Syndrome Group B Protein Prevents the Accumulation of Damaged Mitochondria by Promoting Mitochondrial Autophagy., J Exp Med 209, 855, 2012 |
| 小鼠 | 小鼠 | Epidermal and dermal | Collagenase Type 4: 0.18% | PBS | King, I., Kroenke, M. and Segal, B.: GM-CSF-Dependent, CD103+ Dermal Dendritic Cells Play a Critical Role in Th Effector Cell Differentiation After Subcutaneous Immunization., J Exp Med 207, 953, 2010 |
| Collagenase/Dispase: 0.18% | |||||
| 小鼠 | 小鼠, 2-3 天 | Dermal fibroblasts | Collagenase Type 2: 0.04% | DME | Farina, G., York, M., Di Marzio, M., Collins, C., Meller, S., Homey, B., Rifkin, I., Marshak-Rothstein, A., Radstake, T. and Lafyatis, R.: Poly(I:C) Drives Type I IFN- and TGF尾-Mediated Inflammation and Dermal Fibrosis Simulating Altered Gene Expression in Systemic Sclerosis., J Invest Dermatol 130, 2583-93, 2010 |
| Trypsin: 0.025% | |||||
| 小鼠 | 小鼠 | Dermal | Collagenase Type 3: 0.3% | HBSS | Eidsmo, L., Allan, R., Caminschi, I., Van Rooijen, N., Heath, W. and Carbone, F.:Differential Migration of Epidermal and Dermal Dendritic Cells During Skin Infection., J Immunol 182, 3165, 2009 |
| Deoxyribonuclease I: 0.0005% | |||||
| 小鼠 | 小鼠, 新生的 | Dermal | Collagenase: 0.35% | PBS | Crigler Lauren, Kazhanie Amita, Yoon Tae-Jin, Zakhari Julia, Anders Joanna, Taylor Barbara, Virador VictoriaM: Isolation of a mesenchymal cell population from murine dermis that contains progenitors of multiple cell lineages, FASEB J 21, 2050-63, 2007 |
| Deoxyribonuclease I: see reference | |||||
| 小鼠 | 小鼠 | Dermal fibroblasts | Trypsin: 0.25% | DMEM | Baxter Ruth M, Crowell Thomas P, McCrann Margaret E, Frew Erica M, Gardner Humphrey: Analysis of the tight skin (Tsk1/+) mouse as a model for testing antifibrotic agents,Lab Invest 85, 1199-209, 2005 |
| Collagenase Type 1: 0.25% | |||||
| 小鼠 | 小鼠, 新生的 | Microvascular endothelial | Neutral Protease: 0.005% | DMEM | Cha, S., Talavera, D., Demir, E., Nath, A. and Sierra-Honigmann, M.: A Method of Isolation and Culture of Microvascular Endothelial Cells from Mouse Skin., Microvasc Res 70, 198, 2005 |
| Collagenase Type 1: 4% | |||||
| 小鼠 | 小鼠, male 6-24 week | Skin side population | Collagenase Type 4: 0.2% | PBS | Montanaro F, Liadaki K, Volinski J, Flint A, and Kunkel LM.: Skeletal muscle engraftment potential of adult mouse skin side population cells, Proc Natl Acad Sci U S A 100, 9336, 2003 |
| Neutral Protease: 1.2 u/ml | |||||
| 小鼠 | 小鼠 | Ear epidermal | Trypsin: 0.1% | PBS | Takanami-Ohnishi Yoko, Amano Shinya, Kimura Sadao, Asada Sachie, Utani Atsushi, Maruyama Masumi, Osada Hiroyuki, Tsunoda Hajime, Irukayama-Tomobe Yoko, Goto Katsutoshi, Karin Michael, Sudo Tatsuhiko, Kasuya Yoshitoshi: Essential role of p38 mitogen-activated protein kinase in contact hypersensitivity, J Biol Chem 277, 37896-903, 2002 |
| Collagenase: 0.2% | |||||
| 小鼠 | 小鼠, 3-7 月 | Fibroblasts, mesangial, smooth muscle | Trypsin: 0.25% | DMEM | Bradshaw AD, Francki A, Motamed K, Howe C, Sage EH: Primary mesenchymal cells isolated from SPARC-null mice exhibit altered morphology and rates of proliferation, Mol Biol Cell 10, 1569-79, 1999 |
| Collagenase: see reference | |||||
| Soybean Trypsin Inhibitor: .05% | |||||
| 豬 | 豬, 3-4 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, 2-6 月 | Keratinocytes | Neutral Protease: 0.25% | Dulbecco-Vogt MEM | Regauer S, Compton C: Cultured porcine epithelial grafts: an improved method, J Invest Dermatol 94, 230-4, 1990 |
| 大鼠 | Wistar大鼠, 4-8 天 | Dermal fibroblasts and keratinocytes | Trypsin: 0.25% | Ham's F-12 | Sugihara, H and Toda, S: Primary Tissue Intact and Dissociated Cell Culture,Cell & Tissue Culture: Laboratory Procedures Vol. 1,Doyle, A., Griffiths, J., and Newell, D., John Wiley and Sons, Ltd., 3A:2.1, 1995 |
| 大鼠 | SD大鼠, 雄性, 40-60 天 | Sebaceous | Trypsin: 0.2% | DMEM with FBS | Laurent, S., Mednieks, M., and Rosenfield, R.: Growth of Sebaceous Cells in Monolayer Culture, In Vitro Cell Dev Biol 28, 83, 1992 |
| PBS | |||||
| 大鼠 | 大鼠, 新生的, 產后的12-24 小時 | Fibroblasts | Trypsin: 0.2% | HEPES buffered DMEM | Acheson, A., Barker, P., Alderson, R., Miller, F., and Murphy, R.: Detection of Brain-Derived Neurotrophic Factor-like Activity in Fibroblasts and Schwann Cells: Inhibition by Antibodies to NGF, Neuron 7, 265, 1991 |
| 大鼠 | 大鼠, 白化變種, 一天大, CFN | Keratinocytes | Trypsin: 1% | EBSS | Vaughan, F., Gray, R., and Bernstein, I.: Growth and Differentiation of Primary Rat Keratinocytes on Synthetic Membranes, In Vitro Cell Dev Biol 22 (3), 141, 1986 |
我們公司zui大優勢是強大的采購，
1：基本什么都能進口，血清，抗體，耗材，還有部分限制進口的，
2：貨品全，現經營過700多個品牌，基本所有生物試劑耗材都可以進口，特別是冷偏的產品那就更有優勢，
3：提供加急服務，一般1-2周到貨，超過時限加急費全免
4：價格公道，絕大部分價格有優勢，當然不能保證100%產品都是,因為意味著沒有服務.
5：良好的信譽，大部分客戶我們提供貨到付款服務，客戶包括清華，北大 交大 復旦，中山等100多所大學，ROCHE，阿斯利康，國藥 ，fisher等500多家公司
6：我們還是Santa,Advanced Biotechnologies Inc, Worthington Biochemical ,bangs,BBInternational,crystalchem,dianova,FD Neurotechnologies,Inc. Dako,Inc, GePromegaridege, Glycotope Biotechnology GmbH; iduron,Innovative Research of America, Ludger, neuroprobe,Worthington Biochemical , Polysciences,prospecbi, QA-BIO,quickzyme,FULLERLABS,INC,sterlitech；sysy,TriLink BioTechnologies,Inc；worthington-biochem,zyagen等幾十家國外公司授權代理。
7：我們還是invitrogen,qiagen,Worthington Biochemical am,sigma;Promega,roche,merck, rnd,BD, GE,pierce,BioLegend等*批發，歡迎合作。
