L Wnt-5A細胞

是否是腫瘤細胞： 0

物種來源： 小鼠

生長狀態： 貼壁生長

運輸方式： 凍存運輸

細胞形態： 成纖維樣

數量： 大量

ATCC Number： CRL-2814?

組織來源： subcutaneous connective tissue; areolar and adipose

Designations: L Wnt-5A

Depositors: R Nusse

Biosafety Level: 1

Shipped: frozen

Medium & Serum: See Propagation

L Wnt-5A細胞Growth Properties: adherent

Organism: Mus musculus

Morphology: fibroblast

Source: Tissue: subcutaneous connective tissue; areolar and adipose

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Comments: L-M(TK-) cells (ATCC CCL-1.3) were transfected with a Wnt-5A expression vector and stable clones were selected in medium containing G418. The Wnt-5A gene encodes a secreted glyoprotein with a variety of signaling effects. Wnt proteins bind to receptors of the Frizzled family on the cell surface. Wnt genes control many of the patterning and growth events during embryonic development. The cells secrete biologically active Wnt-5A protein. L Wnt-5A細胞They are presently the best source for production of Wnt-5A conditioned medium. Since the conditioned medium contains other factors besides the Wnt-5A protein, it is necessary to control any experiments involving the Wnt-5A conditioned medium with control conditioned medium from the parental cell line (ATCC CRL-2648).

Propagation: ATCC complete growth medium: The base medium for this cell line is ATCC Hybri-Care Medium, Catalog No. 46-X. Hybri-Care Medium is supplied as a powder and should be reconstituted in 1 L cell-culture-grade water and supplemented with 1.5 g/L sodium bicarbonate. To make the complete growth medium, add the following components to the base medium: fetal bovine serum to a final concentration of 10%.

Atmosphere: air, 95%; carbon dioxide (CO2), 5%

Temperature: 37.0°C

Subculturing: Protocol:

Remove and discard culture medium.

Briefly rinse the cell layer with 0.25% (w/v) Trypsin- 0.53 mM EDTA solution to remove all traces of serum that contains trypsin inhibitor.

L Wnt-5A細胞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 gently pipetting.

Add appropriate aliquots of the cell suspension to new culture vessels.

An inoculum of 5 X 10(3) to 3 X 10(4) viable cells/cm2 is recommended.

Incubate cultures at 37?C.

Protocol for Wnt-5A conditioned medium: 1. Split the cells 1:10 in 10 ml culture medium without G418 in 10 cm petri dishes and let the cells grow for 4 days (approximately to confluency). 2. Take off the medium and sterile filter. This is the first batch of medium. 3. Add 10 ml fresh culture medium without G418 and culture for another 3 days. 4. Take off the medium and sterile filter. This is the second batch of medium. Discard the cells, because they will be overgrown. 5. Mix the first batch and second batch of medium 1:1. This is the Wnt-5A conditioned medium. It is stable at 4 degrees C and can be frozen.

Interval: Maintain cultures at a cell concentration between 7 X 10(3) and 3 X 10(5) cells/cm2

Subcultivation Ratio: L Wnt-5A細胞A subcultivation ratio of 1:4 to 1:6 is recommended

Preservation: Freeze medium: Complete growth medium supplemented with 5% (v/v) DMSO

Storage temperature: liquid nitrogen vapor phase

Doubling Time: 36 hrs

Related Products: Recommended medium (without the additional supplements or serum described under ATCC Medium):ATCC 30-2002

recommended serum:ATCC 30-2020

parental cell line:ATCC CRL-2648

References: 88964: Chen W, et al. Dishevelled 2 recruits beta-arrestin 2 to mediate Wnt5A-stimulated endocytosis of Frizzled 4. Science 301: 1391-1339, 2003. PubMed: 12958364