KC-2701

S180-Epcam-Midddle Cell Line

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Background of S180-Epcam-Midddle Cell Line

Epithelial cell adhesion molecule (EpCAM) is a transmembrane glycoprotein functioned as cell-cell adhesion in epithelia, cell migration, proliferation and differentiation. EpCAM have oncogenic potential through upregulation of c-myc, cyclins A & E after cleavage. Epcam is not only used as diagnostic marker for various cancer, but also a potential target for cancer therapy.

Specifications

Catalog Number	KC-2701
Cell Line Name	S180-Epcam-Midddle Cell Line
Host Cell Line	Mouse S180 cell line
Description	Stable S180 cell line expressing exogenous human Epcam gene in middle level
Quantity	One vial 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 + 2μg/ml Puromycin
Selection Marker	Puromycin
Morphology	Rounded
Subculture	Split saturated culture 1:2-1:3 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 14-20 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

S180-Epcam-Middle cell Line was generated using a retroviral vector expressing human Epcam sequence.

Characterization

Figure 1: Characterization of human Epcam overexpression in the S180-Epcam stable clone using FACS.

Cell Resuscitation

1. Prewarm culture medium (RPMI1640 supplemented with 10% FBS and 2μg/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 1:2-1:3 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. Litvinov, Sergey; et al. (1994). Ep-CAM: a human epithelial antigen is a homophilic cellÿcell adhesion molecule. The Journal of Cell Biology. 125 (2): 437ÿ46. 2. Maetzel, Dorothea; et al. (2009). Nuclear signalling by tumour-associated antigen EpCAM. Nature Cell Biology. 11 (2): 162ÿ71. 3. Osta, WA; et al. (2004). EpCAM is overexpressed in breast cancer and is a potential target for breast cancer gene therapy. Cancer Res. 64 (16): 5818ÿ24.
2. Gaber A, Lenarčič B, Pavšič M. Current View on EpCAM Structural Biology. Cells. 2020 May 31;9(6):1361. doi: 10.3390/cells9061361. PMID: 32486423; PMCID: PMC7349879.

Use License Agreement

Research Use Only.

Not for use in diagnostic procedures or therapeutic applications.

Redistribution of the cell line or its derivatives is prohibited without prior written permission from Kyinno Biotechnology.