KC-2147

B16/F10-CLDN6-Cell-Line

23480

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Background of B16/F10-CLDN6-Cell-Line

CLDN6, also known as claudin 6, is a multi-transmembrane protein in the Claudin family. CLDN6 is expressed by epithelial cells where it participates in tissue development and the maintenance of tight junction integrity. And it is one of the entry cofactors for hepatitis C virus. The methylation of CLDN6 may be involved in esophageal tumorigenesis.

Specifications

Catalog Number	KC-2147
Cell Line Name	B16/F10-CLDN6-Cell-Line
Host Cell Line	Mouse B16F10 cell line
Description	Stable B16F10 clone expressing exogenous human CLDN6 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% McCoy's 5a+20% FBS+10% DMSO
Propagation Medium	McCoy5A+10%FBS+1μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Epithelial
Subculture	Split saturated culture 1:4-1:10 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 17 hours
Mycoplasma Status	Negative
In Vivo Validation	Yes

Cell Line Generation

B16F10 human CLDN6 cell Line was generated using a lentiviral vector expressing human CLDN6 sequence.

Characterization

Cell Resuscitation

Prewarm culture medium (Mycoy5A + 10% FBS + 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 1:4-1:10 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% McCoy's 5a + 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

Morita K, Furuse M, Fujimoto K, Tsukita S (Mar 1999). "Claudin multigene family encoding four-transmembrane domain protein components of tight junction strands". Proc Natl Acad Sci U S A. 96 (2): 511ÿ6. Bibcode:1999PNAS...96..511M. doi:10.1073/pnas.96.2.511. PMC 15167. PMID 9892664.
Meertens L, Bertaux C, Cukierman L, Cormier E, Lavillette D, Cosset FL, Dragic T (Mar 2008). "The Tight Junction Proteins Claudin-1, -6, and -9 Are Entry Cofactors for Hepatitis C Virus". J Virol. 82 (7): 3555ÿ60. doi:10.1128/JVI.01977-07. PMC 2268462. PMID 18234789.
Entrez Gene: CLDN6 claudin 6.
Aberrant expression of claudin-6 contributes to malignant potentials and drug resistance of cervical adenocarcinoma. Ito, Takasawa, Takasawa et al.Cancer Sci (2022) 113 (4), 1519-1530