KC-3976

CHOK1-CD46-high Cell Line

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Background of CHOK1-CD46-high Cell Line

Membrane cofactor protein (MCP; CD46), a ubiquitously expressed complement regulatory protein, CD46 plays roles in human reproduction, autophagy, modulating T cell activation and effector functions and is a member of the newly identified intracellular complement system (complosome). The protein encoded by this gene may be involved in the fusion of the spermatozoa with the oocyte during fertilization. CD46 could be an important link in the action mechanism of ADCC and CDC intercommunication and may be used for the development of novel therapeutic strategies.

Specifications

Catalog Number	KC-3976
Cell Line Name	CHOK1-CD46-high Cell Line
Clone Number	3#
Host Cell Line	CHOK1
Description	Stable CHOK1 cell line high expressing exogenous CD46 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 CD46 cell line was generated using a lentiviral vector expressing the CD46 sequence.

Characterization

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

Do MH, Thanh HD, To PK, Kim MS, Moon C, Jung C. CD46 protects the bladder cancer cells from cetuximab-mediated cytotoxicity. Sci Rep. 2022 Dec 27;12(1):22420.
Sherbenou DW, Aftab BT, Su Y, Behrens CR, Wiita A, Logan AC, Acosta-Alvear D, Hann BC, Walter P, Shuman MA, Wu X, Atkinson JP, Wolf JL, Martin TG, Liu B. Antibody-drug conjugate targeting CD46 eliminates multiple myeloma cells. J Clin Invest. 2016 Dec 1;126(12):4640-4653.
Liszewski MK, Atkinson JP. Membrane cofactor protein (MCP; CD46): deficiency states and pathogen connections. Curr Opin Immunol. 2021 Oct;72:126-134.