KC-2971

CHOK1-CLEC2D Cell Line

26843

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Background of CHOK1-CLEC2D Cell Line

CLEC2D (C-Type Lectin Domain Family 2 Member D) is a Protein Coding gene. This gene encodes a member of the natural killer cell receptor C-type lectin family. The encoded protein inhibits osteoclast formation,Inhibits bone resorption and Modulates the release of interferon-gamma. CLEC2D uses gene splicing to generate protein isoforms that are structurally distinct and that have different biological activities.

Specifications

Catalog Number	KC-2971
Cell Line Name	CHOK1-CLEC2D Cell Line
Host Cell Line	CHOK1
Description	Stable CHOK1 cell line expressing exogenous CLEC2D 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 CLEC2D Cell Line was generated using a lentiviral vector expressing the CLEC2D sequence.

Characterization

Figure 1: Characterization of CLEC2D overexpression in the CHOK1 CLEC2D stable clone using FACS.

Cell Resuscitation

1. Prewarm culture medium (RPMI1640 supplemented with 10% FBS and 10μg/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 1:4-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

1. Prepare the freezing medium (70% RPMI1640 + 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. Germain C, Bihl F, Zahn S, Poupon G, Dumaurier MJ, Rampanarivo HH, Padkjær SB, Spee P, Braud VM. Characterization of alternatively spliced transcript variants of CLEC2D gene. J Biol Chem. 2010 Nov 12;285(46):36207-15.
2. Mathew PA, Chuang SS, Vaidya SV, Kumaresan PR, Boles KS, Pham HT. The LLT1 receptor induces IFN-gamma production by human natural killer cells. Mol Immunol. 2004 Mar;40(16):1157-63.