KC-1828

293T-CCR8 Cell Line

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Background

CCR8, also named as CDw198, is a GPCR protein belonging to the beta chemokine receptor family, and is mainly expressed in the thymus. CCR8 play an important role in regulation of monocyte chemotaxis and thymic cell apoptosis after interaction with its ligand CCL1.

Specifications

Catalog Number:
KC-1828
Cell Line Name:
293T-CCR8 Cell Line
Host Cell Line:
Human HEK293T cell line
Description:
Stable HEK293T cell line expressing exogenous human CCR8 gene
Quantity:
One vial of frozen cells (5×106 per vial)
Stability:
Stable in culture over a minimum of 10 passages
Application:
Drug screening and biological assays
Freeze Medium:
70% DMEM+20% FBS+10% DMSO
Propagation Medium:
DMEM+10%FBS+1µg/mL Puromycin
Selection Marker:
Puromycin
Morphology:
Epithelial
Subculture:
Split saturated culture 1:3-1:6 every 3 days; seed out at about 1-3 × 105 cells/mL
Incubation:
37 °C with 5% CO2
Storage:
Liquid nitrogen immediately upon receiving
Doubling Time:
Approximately 30 hours
Mycoplasma Status:
Negative

Cell Line Generation

293T human CCR8 cell line was generated using a lentiviral vector expressing the human CCR8 sequence.

Characterization

Application

Cell Resuscitation

1. Prewarm culture medium (DMEM + 10% FBS + 1μg/mL puromycin)in a 37°C water bath.
2. Thaw the frozen vial in a 37°C water bath for 1-2 minutes.
3. Transfer the vial into biosafety cabinet, and wipe the surface with 70% ethanol.
4. Unscrew the top of the vial and transfer the cell suspension gently into a sterile centrifuge tube containing 9.0mL complete culture medium.
5. Spin at ~ 125 × g for 5-7 minutes at room temperature, and discard the supernatant without disturbing the pellet.
6. Resuspend cell pellet with the appropriate volume of complete medium and transfer the cell suspension into a T25 culture flask.
7. Incubate the flask at 37°C, 5% CO2 incubator.
8. Split saturated culture 1:3-1:6 every 3 days; seed out at about 1-3 × 105 cells/mL.

Cell Freezing

1. Prepare the freezing medium (70% DMEM + 20% FBS + 10% DMSO) fresh immediately before use.
2. Keep the freezing medium on ice and label cryovials.
3. Transfer cells to a sterile, conical centrifuge tube, and count the cells.
4. Centrifuge the cells at 250×g for 5 minutes at room temperature and carefully aspirate off the medium.
5. Resuspend the cells at a density of at least 3×106 cells/mL in chilled freezing medium.
6. Aliquot 1 mL of the cell suspension into each cryovial.
7. Freeze cells in the CoolCell freezing container overnight in a -80°C freezer.
8. Transfer vials to liquid nitrogen for long-term storage

References

1. Roos RS, Loetscher M, Legler DF, Clark-Lewis I, Baggiolini M, Moser B (1997). "Identification of CCR8, the receptor for the human CC chemokine I-309". J. Biol. Chem. 272 (28): 17251ÿ4.
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