L Wnt-3A 小鼠皮下結締組織細胞信息 由于這種條件培養基還包含Wnt-3A蛋白外的其他因子，對于涉及Wnt-3A條件培養基的實驗有必要用其來源細胞株(ATCC CRL-2648)作對照。L-M(TK-) cells (ATCC CCL-1.3) 用Wnt-3A表達載體轉染并在含G418的培養基中篩選穩轉株。 Wnt-3A基因編碼一個有變異的信號功效分泌性糖蛋白。 Wnt基因控制胚胎發過程中的許多模式形成和生長事件。 這些細胞分泌有生物活性的Wnt-3A蛋白。 它們是目前生產Wnt-3A條件培養基的來源。 動物種別：小鼠 性別：雄 L Wnt-3A 小鼠皮下結締組織 組織來源： 皮下結締組織；疏松結締組織結締組織和脂肪 品系： C3H/An 由于這種條件培養基還包含Wnt-3A蛋白外的其他因子，對于涉及Wnt-3A條件培養基的實驗有必要用其來源細胞株培養條件：DMEM培養基，90%；0.4 mg/ml G-418；胎牛血清，10% 以下是ATCC 的介紹 L Wnt-3A (ATCC® CRL-2647™) Organism Mus musculus, mouse Tissue subcutaneous connective tissue; areolar and adipose Product Format frozen Morphology fibroblast Culture Properties adherent Biosafety Level 1 Age 100 days Gender male Strain C3H/An Applications This cell line is presently the best source for production of Wnt-3A conditioned medium. Storage Conditions liquid nitrogen vapor phase
Derivation L Wnt-3A 小鼠皮下結締組織 L-M(TK-) cells (ATCC CCL-1.3) were transfected with a Wnt-3A expression vector and stable clones were selected in medium containing G418. Clinical Data male Genes Expressed Wnt-3A Comments The Wnt-3A gene encodes a secreted glyoprotein with a variety of signaling effects. Wnt genes control many of the patterning and growth events during embryonic development.
The cells secrete biologically active Wnt-3A protein. They are presently the best source for production of Wnt-3A conditioned medium.
Since the conditioned medium contains other factors besides the Wnt-3A protein, it is necessary to control any experiments involving the Wnt-3A conditioned medium with control conditioned medium from the parental cell line (ATCC CRL-2648). L Wnt-3A 小鼠皮下結締組織
Complete Growth Medium The base medium for this cell line is ATCC-formulated Dulbecco's Modified Eagle's Medium, Catalog No. 30-2002. To make the complete growth medium, add the following components to the base medium: 0.4 mg/ml G-418; fetal bovine serum to a final concentration of 10%. Subculturing Volumes used in this protocol are for 75 cm2 flask; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes.
Remove and discard culture medium. Briefly rinse the cell layer with 0.25% (w) Trypsin-0.53 mM EDTA solution to remove all traces of serum that contains trypsin inhibitor. Add 2.0 to 3.0 mL of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 5 to 15 minutes). Note: To avoid clumping, do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Cells that are difficult to detach may be placed at 37°C to facilitate dispersal. Add 6.0 to 8.0 mL of complete growth medium and aspirate cells by pipetting gently. Add appropriate aliquots of cell suspension to new culture vessels. Incubate culture vessels at 37°C.
Protocol for Wnt-3A Conditioned Medium: L Wnt-3A 小鼠皮下結締組織 Split the cells 1:10 in 10 mL culture medium (without G418 if conditioned medium is to be used with a cell line sensitive to G418) in 10 cm tissue culture dishes or T-75 flasks and let the cells grow for 4 days (approximay to confluency). Take off the medium and sterile filter. This is the first batch of medium. Add 10 mL fresh culture medium and culture for another 3 days. Take off the medium and sterile filter. This is the second batch of medium. Discard the cells, because they will be overgrown. Mix the first batch and second batch of medium 1:1. This is the Wnt-3A conditioned medium. It is stable at 4°C and can be frozen.
Subc*tion Ratio: A subc*tion ratio of 1:4 to 1:10 is recommended. Medium Renewal: Every 2 to 3 days. Note: For more information on enzymatic dissociation and subculturing of cell lines consult Chapter 10 in Culture of Animal Cells, a manual of Basic Technique by R. Ian Freshney, 3rd edition, published by Alan R. Liss, N.Y., 1994. L Wnt-3A 小鼠皮下結締組織 Cryopreservation Freeze medium: Complete growth medium supplemented with 5% (v) DMSO Storage temperature: liquid nitrogen vapor phase Culture Conditions Atmosphere: air, 95%; carbon dioxide (CO2), 5% Temperature: 37°C
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) Trypsin-0.53 mM EDTA solution to remove all traces of serum that contains trypsin inhibitor.
