KC-0221

293T-CD30-Cell-Line

20041

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Background of 293T-CD30-Cell-Line

CD30, also named as Ki-1 and TNFRSF8, is a transmembrane protein of TNF receptor superfamily. CD30 is mainly expressed on antigen-stimulated Th cells and B cells in physiological condition, play a key role in antigen-induced Th cell proliferation and cytokine secretion. CD30 is also highly expressed in Hodgkin’s disease and others lymphoma, is a biomarker and drug target, its monoclonal antibody Brentuximab had been approved for tumor treatment.

Specifications

Catalog Number	KC-0221
Cell Line Name	293T-CD30-Cell-Line
Host Cell Line	Human HEK293T cell line
Description	HEK293T cell line stable expressing exogenous human CD30 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+0.5µg/mL Puromycin
Selection Marker	Puromycin
Morphology	Epithelial
Subculture	Split saturated culture 1:3-1:6 every 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 human CD30 cell line was generated using lentiviral vector expressing human CD30 sequence.

Characterization

Figure: Characterization of CD30 overexpressing in 293T stable clones using FACS.

Cell Resuscitation

Prewarm culture medium (DMEM + 10% FBS + 0.5μg/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 1:3-1:6 every 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

Molecular cloning and expression of a new member of the nerve growth factor receptor family that is characteristic for Hodgkin's disease. D°îrkop H et al. Cell 1992 Feb;68(3)421-427