| Field | Specification |
|---|---|
| Mfr No | |
| Species |
The SVI cell line has been cloned from the outgrowth of glomeruli which were isolated from H-2kb-tsA58 transgenic mice. The mice carry a temperature-sensitive variant of the SV40 large T antigen under control of the IFN-g-inducible H-2kb promoter. Cells proliferate at 33 degree Celsius, and they differentiate at 37 degree Celsius. At present, the cells have been cultured successfully for more than 40 passages without noting phenotypic changes. SVI are very similar to E11 in terms of morphology and the expression of several markers. For example, podocin and WT1 are expressed to a lesser extent as compared to E11. Differentiation: Start the differentiation process by placing the non-confluent flask(s) into an incubator at 38 degree Celsius / 5% CO2 for a minimum of 14 days to complete the differentiation. Addition of interferon-gamma (INF-gamma) is not necessary.
—
Protein expression: WT1, Lmx1b, nephrin, NEPHI, FAT, P-cadherin, CD2AP, ZO-I, podocalyxin, podoplanin, synpo, podocin, TRPC6 and GAPDH.
- cultureMedium: RPMI 1640, w: 2.0 mM stable Glutamine, w: 2.0 g/L NaHCO3 (Cytion article number 820700a)
- supplements: Supplement the medium with 10% FBS
- dissociationReagent: Accutase
- subculturing: Remove the old medium from the adherent cells and wash them with PBS that lacks calcium and magnesium. For T25 flasks, use 3-5 ml of PBS, and for T75 flasks, use 5-10 ml. Then, cover the cells completely with Accutase, using 1-2 ml for T25 flasks and 2.5 ml for T75 flasks. Let the cells incubate at room temperature for 8-10 minutes to detach them. After incubation, gently mix the cells with 10 ml of medium to resuspend them, then centrifuge at 300xg for 3 minutes. Discard the supernatant, resuspend the cells in fresh medium, and transfer them into new flasks that already contain fresh medium.
- seedingDensity: Inocculate T75 cell culture flasks with 1x 104 cells/cm2 (about 60.000 cells/ml, 12ml medium in one T75) for the proliferation process. Keep the cells at 33 degree Celsius / 5% CO2, until the flask is about 75% confluent.
- fluidRenewal: 3 times per week
- freezeMedium: As a cryopreservation medium, use complete growth medium (including FBS) + 10% DMSO for adequate post-thaw viability, or CM-1 (Cytion catalog number 800100), which includes optimized osmoprotectants and metabolic stabilizers to enhance recovery and reduce cryo-induced stress.
10%
OFF
15%OFF
20%
OFF
$50
OFF
FREE
SAMPLE
