KC-2483

293T-cyno-CD3E-CD3D-CD3G-Cell-Line

24152

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Background of 293T-cyno-CD3E-CD3D-CD3G-Cell-Line

CD3E (CD3 Epsilon Subunit Of T-Cell Receptor Complex) is a protein coding gene.CD3E encodes a protein that is part of the T-cell receptor/CD3 complex (TCR/CD3 complex) and is involved in T-cell development and signalling. Associated pathways include NFAT and TCR signalling in the immune response.

Specifications

Catalog Number	KC-2483
Cell Line Name	293T-cyno-CD3E-CD3D-CD3G-Cell-Line
Host Cell Line	293T
Description	Stable 293T clone expressing exogenous cyno CD3D, CD3E and CD3G 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% DMEM + 20% FBS + 10% DMSO
Propagation Medium	DMEM + 10% FBS + 1μg/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

293T cyno-CD3E-CD3D-CD3G cell line was generated using a lentiviral vector expressing the cyno CD3D, CD3E and CD3G sequence.

Characterization

Figure 1: Characterization of cyno-CD3 overexpression in the 293T cyno-CD3E-CD3D-CD3G stable clone using FACS.

Cell Resuscitation

Prewarm culture medium (DMEM supplemented with 10% FBS and 1μg/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% DMEM + 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

Structure of the parathyroid hormone receptor C terminus bound to the G-protein dimer Gbeta1gamma2. PMID: 18611381 PMCID: PMC2601695 DOI: 10.1016/j.str.2008.04.010.
Zhang Q, Li H, Gao S, Wang J, Li C, Shu J, Lin J. CD3E as a new predictive biomarker of response to omalizumab treatment in asthma patients: Evidence from bioinformatic analysis. Int Immunopharmacol. 2021 Apr;93:107423. doi: 1016/j.intimp.2021.107423. Epub 2021 Feb 10. PMID: 33578181.