KC-6431

CT26-mouse-CLDN18.2 Cell Line

62897

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Background of CT26-mouse-CLDN18.2 Cell Line

CLDN18.2 encodes a member of the claudin family. Claudins are integral membrane proteins and components of tight junction strands. Tight junction strands serve as a physical barrier to prevent solutes and water from passing freely through the paracellular space between epithelial or endothelial cell sheets, and also play critical roles in maintaining cell polarity and signal transductions. This gene is upregulated in patients with ulcerative colitis and highly overexpressed in infiltrating ductal adenocarcinomas. PKC/MAPK/AP-1 dependent pathway regulates the expression of this gene in gastric cells. Alternatively spliced transcript variants encoding different isoforms have been identified.

Specifications

Catalog Number	KC-6431
Cell Line Name	CT26-mouse-CLDN18.2 Cell Line
NCBI/UniProt Accession Number	NM_001194921
Clone Number	3#
Host Cell Line	CT26
Description	CT26 cell line stably expressing exogenous mouse-CLDN18.2 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 + 10 μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Epithelial
Subculture	Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-2 × 10⁵ cells/mL
Incubation	37 °C with 5% CO₂
Storage	Liquid nitrogen immediately upon receiving
Doubling Time	Approximately 30 hours
Mycoplasma Status	Negative

Cell Line Generation

CT26-mouse-CLDN18.2 cell line was generated using a lentiviral vector expressing the mouse-CLDN18.2 sequence.

Characterization

Figure 1: Characterization of mouse-CLDN18.2 overexpression in CT26-mouse-CLDN18.2 stable clones using FACS.

Figure 2: Characterization of CT26-mouse-CLDN18.2 cell line stable clone using PCR sequencing.

Cell Resuscitation

Prewarm culture medium (RPMI1640 supplemented with 10% FBS and 10 μg/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 9.0 mL 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 1-2 × 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

Hayashi D, Tamura A, Tanaka H, Yamazaki Y, Watanabe S, Suzuki K, Suzuki K, Sentani K, Yasui W, Rakugi H, Isaka Y, Tsukita S. Deficiency of claudin-18 causes paracellular H+ leakage, up-regulation of interleukin-1β, and atrophic gastritis in mice. Gastroenterology. 2012 Feb;142(2):292-304. doi: 10.1053/j.gastro.2011.10.040. Epub 2011 Nov 10. PMID: 22079592.