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1、Discovery Labware, Inc., Two Oak Park, Bedford, MA 01730, Tel: 1.978.442.2200 (U.S.) CLSTechServC For Research Use Only. Not for use in diagnostic or therapeutic procedures.For a listing of trademarks, visit 2013 Corning Incorporated SPC-354248 Rev 2.0 - 1 - GUIDELINES FOR USE PRODUCT: Corning? Ma
2、trigel? Basement Membrane Matrix High Concentration, 10 ml vial CATALOG NUMBER:354248 BACKGROUND:Basement membranes are thin extracellular matrices underlying cells in vivo. Corning Matrigel Matrix High Concentration (HC) is a solubilized basement membrane preparation extracted from the Engelbreth-H
3、olm-Swarm (EHS) mouse sarcoma, a tumor rich in extracellular matrix proteins. Its major component is laminin, followed by collagen IV, heparan sulfate proteoglycans, entactin/nidogen.1,2 Corning Matrigel Matrix HC also contains TGF-beta, epidermal growth factor, insulin-like growth factor, fibroblas
4、t growth factor, tissue plasminogen activator,3,4 and other growth factors which occur naturally in the EHS tumor. Corning Matrigel Matrix HC is effective for the attachment and differentiation of both normal and transformed anchorage dependent epithelioid and other cell types. These include neurons
5、,5,6hepatocytes,7 Sertoli cells,8,9 chick lens,10 and vascular endothelial cells.11 Corning Matrigel Matrix HC will influence gene expression in adult rat hepatocytes12,13 as well as three dimensional culture in mouse14-17 and human18,19 mammary epithelial cells. It is the basis for several types of
6、 tumor cell invasion assays,20,21 will support in vivoperipheral nerve regeneration,22-24 and provides the substrate necessary for the study of angiogenesis both in vitro25,26 and in vivo.27-29 Corning Matrigel Matrix HC also supports in vivo propagation of human tumors in immunosupressed mice.30-32
7、 For further information, go to our website at SOURCE:Engelbreth-Holm-Swarm (EHS) Mouse Tumor FORMULATION:Dulbeccos Modified Eagles Medium with 50 g/ml gentamycin Corning Matrigel Matrix HC is compatible with all culture media STABILITY:Stable for a minimum of three months from day of shipment when
8、 stored at -20 CKEEP FROZEN RECONSTITUTIONAND USE:Color variations may occur in frozen or thawed vials of Corning Matrigel Matrix HC, ranging from straw yellow to dark red due to the interaction of carbon dioxide with the bicarbonate buffer and phenol red. Variation in color is normal, does not affe
9、ct product efficacy, and will disappear upon equilibration with 5% CO2. Once Corning Matrigel Matrix HC is thawed, swirl vial to be sure that material is evenly dispersed. Handle using sterile technique. Place thawed vial of Corning Matrigel Matrix HC in sterile area, spray top of vial with 70% ETOH
10、 and air dry. Corning Matrigel Matrix HC may be gently pipetted using a pre-cooled pipette to ensure homogeneity. Corning Matrigel Matrix HC may be used as a thin gel layer (0.5mm), with cells plated on top. Cells may also be cultured inside the Corning Matrigel Matrix HC, using a 1 mm layer. Extens
11、ive dilution will result in a thin, non-gelled protein layer. This may be useful for cell attachment, but may not be as effective in differentiation studies. Corning Matrigel Matrix HC can be used to assess in vivo angiogenic activity of different compounds by subcutaneous injection into mice (Corni
12、ng Matrigel Plug Assay).2,8,25 The high protein 名师资料总结 - - -精品资料欢迎下载 - - - - - - - - - - - - - - - - - - 名师精心整理 - - - - - - - 第 1 页，共 3 页 - - - - - - - - - Discovery Labware, Inc., Two Oak Park, Bedford, MA 01730, Tel: 1.978.442.2200 (U.S.) CLSTechServC For Research Use Only. Not for use in diagnos
13、tic or therapeutic procedures.For a listing of trademarks, visit 2013 Corning Incorporated SPC-354248 Rev 2.0 - 2 - concentration augments the growth of tumors and also allows the Corning? Matrigel?Plug to maintain its integrity after injection. This keeps the injected tumor and/or angiogenic compo
14、unds localized for in situ analysis and/or future excision. Dispense remaining material into appropriate aliquots, using pre-cooled tubes, and refreeze immediately. Avoid multiple freeze thaws. DO NOT STORE IN FROST-FREE FREEZER.CAUTION : Corning Matrigel Matrix HC will gel rapidly at 22oC to 35oC.
15、Thaw overnight at 4oC on ice (Matrigel may gel at slightly elevated temperatures in a refrigerator). Keep product on ice before use, and use pre-cooled pipettes, tips, and tubes when preparing Corning Matrigel Matrix HC for use. Gelled Corning Matrigel Matrix HC may be re-liquified if placed at 4C o
16、n ice for 24-48 hours. INJECTION PROTOCOL: 1.It is critical to keep the Corning Matrigel Matrix HC and the Corning Matrigel/Cell suspension as cold as possible, without freezing, prior to injecting into the mice. It is very important to keep the Corning Matrigel and the Corning Matrigel/Cell suspens
17、ion as asceptic as possible throughout the procedure. 2.For each recipient mouse, mix cells (2 x 105 or greater) and Corning Matrigel Matrix HC together in a final volume of 0.5 ml on ice. 3.The cells should be in as small a volume as possible. Typically, 250 l ice cold medium containing 2 x 106cell
18、s/ml is mixed with 250 l ice cold Corning Matrigel Matrix HC. 4.Inject the cells subcutaneously in athymic mice using a 19G needle for tissue samples and a 23G needle for cultured cells. The injections should be done quickly to prevent the Matrigel from solidifying. 5.Rotate the syringe when withdra
19、wing to prevent leakage. The needles will need to be changed frequently due to blockage. NOTE: For more details on this application go to to access CLS-DL-CC-036 (Technical Bulletin 455: Methods for Implantation of Corning Matrigel Matrix into Mice and Tissue Fixation). CELL RECOVERY:Dispase (Catal
20、og No. 354235), Corning Cell Recovery Solution (Catalog No. 354253) Most efficient recovery of cells growing on Corning Matrigel Matrix HC is accomplished using Corning Cell Recovery Solution that depolymerizes the Matrigel Matrix within 7 hours on ice or with Dispase, a metalloenzyme which gently r
21、eleases the cells allowing for continuous culture. REFERENCES: 1. Kleinman, H.K., et al., Isolation and characterization of type IV procollagen, laminin, and heparan sulfate proteoglycan from the EHS sarcoma, Biochemistry, 21:6188 (1982). 2. Kleinman, H.K., et al., Basement membrane complexes with b
22、iological activity, Biochemistry, 25:312 (1986). 3. Vukicevic, S., et al., Identification of multiple active growth factors in basement membrane Matrigel suggests caution in interpretation of cellular activity related to extracellular activity related to extracellular matrix components, Experimental
23、 Cell Research, 202:1 (1992). 4. McGuire, P.G. and Seeds, N.W., The interaction of plasminogen activator with a reconstituted basement membrane matrix and extracellular macromolecules produced by cultured epithelial cells, J. Cell. Biochem., 40:215 (1989). 名师资料总结 - - -精品资料欢迎下载 - - - - - - - - - - -
24、- - - - - - - 名师精心整理 - - - - - - - 第 2 页，共 3 页 - - - - - - - - - Discovery Labware, Inc., Two Oak Park, Bedford, MA 01730, Tel: 1.978.442.2200 (U.S.) CLSTechServC For Research Use Only. Not for use in diagnostic or therapeutic procedures.For a listing of trademarks, visit 2013 Corning Incorporated
25、 SPC-354248 Rev 2.0 - 3 - 5. Biederer, T. and Scheiffele, P., Mixed-culture assays for analyzing neuronal synapse formation, Nature Protocols, 2(3):670 (2007). 6. Li, Y., et al., Essential Role of TRPC channels in the guidance of nerve growth cones by brain-derived neurotrophic factor, Nature, 434:8
26、94 (2005). 7. Bi, Y., et al., Use of cryopreserved human hepatocytes in sandwich culture to measure hepatobiliary transport, Drug Metabo. and Dispos., 34(9):1658 (2006). 8. Hadley, M.A., et al., Extracellular matrix regulates sertoli cell differentiation, testicular cord formation, and germ cell dev
27、elopment in vitro, J. Cell Biol., 101:1511 (1985). 9. Yu, X., et al., Essential role of extracellular matrix (ECM) overlay in establishing the functional integrity of primary neonatal rat sertoli cell/gonocyte co-cultures: An improved in vitro model for assessment of male reproductive toxicity, Toxi
28、logical Sciences, 84(2):378 (2005). 10. Ireland, M.E., Quantification and regulation of mRNAs encoding beaded filament proteins in the chick lens, 16(8):838 (1997). 11. McGuire, P.G., and Orkin, R.W., A simple procedure to culture and passage endothelial cells from large vessels of small animals, Bi
29、otechniques, 5(6):456 (1987). 12. Bissel, D.M., et al., Support of cultured hepatocytes by a laminin-rich gel. Evidence for a functionally significant subendothelial matrix in normal rat liver, J. Clinical Invest., 79:801 (1987). 13. Page, J.L., et al., Gene expression profiling of extracellular mat
30、rix as an effector of human hepatocyte phenotype in primary cell culture, Toxilogical Sciences, 97(2):384 (2007). 14 Li, M.L., et al., Influence of a reconstituted basement membrane and its components on casein gene expression and secretion in mouse mammary epithelial cells, Proc. Nat. Acad. Sci. US
31、A, 84:136 (1987). 15 Barcellof, M.H., et al., Functional differentiation and aveolar morphogenesis of primary mammary cultures on reconstituted basement membrane, Development, 105:223 (1989). 16. Roskelley, C.D., et al., Extracellular matrix-dependent tissue-specific gene expression in mammary epith
32、elial cells requires both physical and biochemical signal transduction, Proc. Nat. Acad. Sci. USA, 91(26):12378 (1994). 17. Xu, R., et al., Extracellular matrix-regulated gene expression requires cooperation of SWI/SNF and transcription factors, J. Biol. Chem., 282(20):14992 (2007). 18. Debnath, J.,
33、 et al., Morphogenesis and oncogenesis of MCF-10A mammary epithelial acini grown in three-dimensional basement membrane cultures, Methods, 30(3):256 (2003). 19. Muthuswamy, S.K., et al., ErbB2, but not ErbB1, reinitiates proliferation and induces luminal repopulation in epithelial acini, Nat. Cell B
34、iol., 3(9):785 (2001). 20. Terranova, V.P., et al., Use of a reconstituted basement membrane to measure cell invasiveness and select for highly invasive tumor cells, Proc. Nat. Acad. Sci. USA, 83:465 (1986). 21. Albini, A., et al., A rapid in vitro assay for quantitating the invasive potential of tu
35、mor cells, Cancer Research, 47:3239 (1987). 22. Madison, R., et al., Increased rate of peripheral nerve regeneration using bioresorbable nerve guides and laminin containing gel, Exp. Neurology, 88:767 (1985). 23. Xu, X.M., et al., Axonal regeneration into Schwann cell-seeded guidance channels grafte
36、d into transected adult rat spinal cord, J. Comp. Neurol., 351(1):145 (1994). 24. Fouad, K., et al., Combining schwann cell bridges and olfactory-ensheathing glia grafts with chondroitinase promotes locomotor recovery after complete transection of the spinal cord, The Journal of Neuroscience, 25(5):
37、1169 (2005). 25. Kubota, Y., et al., Role of laminin and basement membrane in the morphological differentiation of human endothelial cells into capillary-like structures, J. Cell Biol., 107:1589 (1988). 26. Maeshima, Y., et al., Identification of the anti-angiogenic site within vascular basement mem
38、brane-derived Tumstatin, J. Biol. Chem., 276(18):15240 (2001). 27. Passaniti, A., et al., A simple, quantitative method for assessing angiogenesis and anti-angiogenic agents using reconstituted basement membrane, heparin, and fibroblast growth factor , Lab Invest., 67:519 (1992). 28. Isaji, M., et a
39、l., Tranilast inhibits the proliferation, chemotaxis and tube formation of human microvascular endothelial cells in vitro and angiogenesis in vivo, British Journal of Pharmacology, 122:1061 (1997). 29. Kisucka, J., et al., Platelets and platelet adhesion support angiogenesis while preventing excessi
40、ve hemorrhage, Proc. Nat. Acad. Sci. USA, 103(4):855 (2006). 30. Albini, A., et al., Matrigel promotes retinoblastoma cell growth in vitro and in vivo, Int. J. Cancer, 52(2):234 (1992). 31. Yue, W., et al., MCF-7 human breast carcinomas in nude mice as a model for evaluating aromatase inhibitors, J.
41、 Steroid Biochem. Molec. Biol., 44(4-6):671 (1993). 32. Angelucci, A., et al., Suppression of EGF-R signaling reduces the incidence of prostate cancer metastasis in nude mice, Endocrine-Related Cancer, 13(1):197 (2006). CALIFORNIA PROPOSITION 65 NOTICE WARNING: This product contains a chemical known to the state of California to cause cancer. Component: Chloroform 名师资料总结 - - -精品资料欢迎下载 - - - - - - - - - - - - - - - - - - 名师精心整理 - - - - - - - 第 3 页，共 3 页 - - - - - - - - -
