KC-2314

293T-cyno-TEK-Cell-Line

23814

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Background of 293T-cyno-TEK-Cell-Line

This gene encodes a receptor that belongs to the protein tyrosine kinase Tie2 family. The encoded protein possesses a unique extracellular region that contains two immunoglobulin-like domains, three epidermal growth factor (EGF)-like domains and three fibronectin type III repeats. The ligand angiopoietin-1 binds to this receptor and mediates a signaling pathway that functions in embryonic vascular development. Mutations in this gene are associated with inherited venous malformations of the skin and mucous membranes. Alternative splicing results in multiple transcript variants. Additional alternatively spliced transcript variants of this gene have been described, but their full-length nature is not known.

Specifications

Catalog Number	KC-2314
Cell Line Name	293T-cyno-TEK-Cell-Line
Host Cell Line	293T
Description	Stable HEK293T clone expressing exogenous cyno TEK 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 TEK cell line was generated using a lentiviral vector expressing the cyno TEK sequence.

Characterization

Figure 1: Characterization of cyno TEK overexpression in the 293T cyno-TEK stable clone using qPCR.

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

Du S, Qian J, Tan S, Li W, Liu P, Zhao J, Zeng Y, Xu L, Wang Z, Cai J. Tumor cell-derived exosomes deliver TIE2 protein to macrophages to promote angiogenesis in cervical cancer. Cancer Lett. 2022 Mar 31;529:168-179.
Jo G, Bae J, Hong HJ, Han AR, Kim DK, Hong SP, Kim JA, Lee S, Koh GY, Kim HM. Structural insights into the clustering and activation of Tie2 receptor mediated by Tie2 agonistic antibody. Nat Commun. 2021 Nov 1;12(1):6287.
Reijrink M, van Ark J, Lexis CPH, Visser LM, Lodewijk ME, van der Horst ICC, Zeebregts CJ, van Goor H, de Jager SCA, Pasterkamp G, Wolffenbuttel BHR, Hillebrands JL. Increased frequency of proangiogenic tunica intima endothelial kinase 2 (Tie2) expressing monocytes in individuals with type 2 diabetes mellitus. Cardiovasc Diabetol. 2022 May 12;21(1):72.