KC-1075

CHOK1-CLEC9A-Cell-Line

21489

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Background of CHOK1-CLEC9A-Cell-Line

CLEC9A（C-Type Lectin Domain Family 9 Member A）， Also known as DNGR-1 (Dendritic Cell NK Lectin Group Receptor-1), it is a receptor protein primarily expressed on the surface of a specific subset of dendritic cells. Its core function is to specifically recognize and bind to dead or damaged cell debris, and initiate immune responses against internal antigens of these cells, playing a key role as a "guardian" and "alarm" in cross presentation and immune surveillance.

Specifications

Catalog Number	KC-1075
Cell Line Name	CHOK1-CLEC9A-Cell-Line
Host Cell Line	CHOK1
Description	Stable CHOK1 clone expressing exogenous CLEC9A 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 1:6-1:8 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-CLEC9A-cell-line was generated using a lentiviral vector expressing the CLEC9A sequence.

Characterization

Figure 1: Characterization of CLEC9A overexpression in the CHOK1-CLEC9A 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 1:6-1:8 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.Hussain Z, Zhang Y, Qiu L, Gou S, Liu K. Exploring Clec9a in dendritic cell-based tumor immunotherapy for molecular insights and therapeutic potentials. NPJ Vaccines. 2025 Feb 7;10(1):27. doi: 10.1038/s41541-025-01084-2. PMID: 39920156; PMCID: PMC11806010.
2.Lahoud MH, Radford KJ. Enhancing the immunogenicity of cancer vaccines by harnessing CLEC9A. Hum Vaccin Immunother. 2022 Dec 31;18(1):1873056. doi: 10.1080/21645515.2021.1873056. Epub 2021 Feb 24. PMID: 33625943; PMCID: PMC8920153.
3.Cheng AC, Yang KY, Chen NJ, Hsu TL, Jou R, Hsieh SL, Tseng PH. CLEC9A modulates macrophage-mediated neutrophil recruitment in response to heat-killed Mycobacterium tuberculosis H37Ra. PLoS One. 2017 Oct 24;12(10):e0186780. doi: 10.1371/journal.pone.0186780. PMID: 29065139; PMCID: PMC5655532.