KC-3363

CHOK1-Rabbit-CCR8 Cell Line

34352

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Background of CHOK1-Rabbit-CCR8 Cell Line

CCR8, also known as CD198, is a member of the chemokine receptor subfamily and belongs to the G-protein coupled receptor, which is expressed predominantly in lymphoid tissues and has also been found in glomerular podocytes and human umbilical vein endothelial cells (HUVECs). CCR8 is associated with Th2 lymphocytes, which are critical for allergy, and has a role in lymphocyte activation, migration, proliferation and differentiation and in allergic diseases. CCR8 may contribute to the proper positioning of activated T cells within the antigenic challenge sites and specialized areas of lymphoid tissues.

Specifications

Catalog Number	KC-3363
Cell Line Name	CHOK1-Rabbit-CCR8 Cell Line
Clone Number	3#
Host Cell Line	CHOK1
Description	Stable CHOK1 cell line expressing exogenous Rabbit CCR8 gene
Quantity	Two vials of frozen cells (≥2-10⁶/vial)
Stability	Stable in culture over a minimum of 10 passages
Application	Drug screening and biological assays
Freezing Medium	70% RPMI1640 + 20% FBS + 10% DMSO
Propagation Medium	RPMI1640 + 10% FBS + 10μg/ml Puromycin
Selection Marker	Puromycin
Morphology	Epithelial
Subculture	Split saturated culture 1:4-1:8 every 2-3 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
In Vivo Validation	NA

Cell Line Generation

CHOK1-Rabbit-CCR8 cell line was generated using a lentiviral vector expressing the Rabbit CCR8 sequence.

Characterization

Figure 1: Characterization of Rabbit CCR8 overexpression in the CHOK1-Rabbit-CCR8 stable clone using FACS.

Cell Resuscitation

Prewarm culture medium (RPMI1640 supplemented with 10% FBS and 10μg/mL Puromycin)in a 37°C water bath.
Thaw the frozen vial in a 37°C water bath for 1-2 minutes.
Transfer the vial into biosafety cabinet, and wipe the surface with 70% ethanol.
Unscrew the top of the vial and transfer the cell suspension gently into a sterile centrifuge tube containing 9.0mL complete culture medium.
Spin at ~ 125 × g for 5-7 minutes at room temperature, and discard the supernatant without disturbing the pellet.
Resuspend cell pellet with the appropriate volume of complete medium and transfer the cell suspension into a T25 culture flask.
Incubate the flask at 37°C, 5% CO₂ incubator.
Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% RPMI1640 + 20% FBS + 10% DMSO) fresh immediately before use.
Keep the freezing medium on ice and label cryovials.
Transfer cells to a sterile, conical centrifuge tube, and count the cells.
Centrifuge the cells at 250×g for 5 minutes at room temperature and carefully aspirate off the medium.
Resuspend the cells at a density of at least 3×10⁶ cells/mL in chilled freezing medium.
Aliquot 1 mL of the cell suspension into each cryovial.
Freeze cells in the CoolCell freezing container overnight in a -80°C freezer.
Transfer vials to liquid nitrogen for long-term storage.

References

Islam SA, Chang DS, Colvin RA, Byrne MH, McCully ML, Moser B, Lira SA, Charo IF, Luster AD (2011). Mouse CCL8, a CCR8 agonist, promotes atopic dermatitis by recruiting IL-5+ T(H)2 cells. Nat. Immunol. 12 (2): 16777.
Moser B. Chemokine Receptor-Targeted Therapies: Special Case for CCR8. Cancers (Basel). 2022 Jan20:14(3):511.
Garlisi CG, Xiao H, Tian F, Hedrick JA, Billah MM, Egan RW, Umland SP (October 1999). The assignment of chemokine-chemokine receptor pairs: TARC and MIP-1 beta are not ligands for human CC-chemokine receptor8.Eur.J.Immunol.29(10):3210-5.