3T3 MEFs KO細(xì)胞

細(xì)胞類型： 其他細(xì)胞類型

是否是腫瘤細(xì)胞： 0

物種來(lái)源： 小鼠

器官來(lái)源： 胚胎

年限： 13.5 gestation embryo

生長(zhǎng)狀態(tài)： 貼壁生長(zhǎng)

運(yùn)輸方式： 凍存運(yùn)輸

數(shù)量： 大量

細(xì)胞形態(tài)： 成纖維樣

ATCC Number： CRL-2753?

3T3 MEFs KO細(xì)胞Designations: 3T3 MEFs KO

Depositors: MP Lisanti

Biosafety Level: 1

Shipped: frozen

Medium & Serum: See Propagation

Growth Properties: adherent

Organism: Mus musculus

Morphology: fibroblast

Source: Organ: embryo

Cell Type: fibroblast fibroblast; spontanous immortalization (3T3)

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Age: 13.5 gestation embryo

Comments: 3T3 MEFs KO細(xì)胞Mice homozygous null for the caveolin-1 gene, Cav-1 (-/-), and their wild-type littermates, Cav-1 (+/+), were generated by targeted disruption of the caveolin-1 gene. A construct was introduced into WW6 embryonic stem (ES) cells by electroporation to disrupt the Cav-1 locus. Mouse embryonic fibroblasts (MEFs) were obtained from day 13.5 littermate mouse embryos and immortalized using the 3T3 protocol [PubMed: 11457855]. The 3T3 MEFs KO cell line (ATCC CRL-2753) is homozygous for a disruption of the caveolin-1 gene Cav-1 (-/-) while the 3T3 MEFs WT cell line (ATCC CRL-2752) is Cav-1 (+/+).Analysis of cultured fibroblasts from Cav-1 null embryos reveals a loss of caveolin-2 protein expression; defects in the endocytosis of a known caveolar ligand, (fluorescein isothiocyanate-albumin); and a hyperproliferative phenotype. These phenotypic changes are reversed by recombinant expression of the caveolin-1 cDNA [PubMed: 11457855]. These cell lines are useful in studying the role of Caveolin-1 in a variety of signaling and membrane trafficking events. A culture deposited with the ATCC in September of 2002 was found to be contaminated with mycoplasma. Progeny were cured by a 21-day treatment with BM Cyclin. The cells were assayed for mycoplasma, by the Hoechst stain, PCR and the standard culture test, after a six-week period following treatment. All tests were negative.

Propagation: ATCC 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: fetal bovine serum to a final concentration of 10%.

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

Temperature: 37.0°C

Subculturing: 3T3 MEFs KO細(xì)胞Protocol: Subculture at 80% confluency.

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.

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.

Incubate cultures at 37?C.

Subcultivation Ratio: A subcultivation ratio of 1:5 to 1:10 is recommended

Medium Renewal: Every 2 to 3 days

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

Storage temperature: liquid nitrogen vapor phase

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

recommended serum:ATCC 30-2020

References: 60275: Razani B, et al. 3T3 MEFs KO細(xì)胞Caveolin-1 null mice are viable but show evidence of hyperproliferative and vascular abnormalities. J. Biol. Chem. 276: 38121-38138, 2001. PubMed: 11457855

60276: Sotgia F, et al. Intracellular retention of glycosylphosphatidyl inositol-linked proteins in caveolin-deficient cells. Mol. Cell. Biol. 22: 3905-3926, 2002. PubMed: 11997523