KC-4179

293T LRP5 Cell Line

43749

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Background of 293T LRP5 Cell Line

LRP5 encodes a transmembrane low-density lipoprotein receptor that binds and internalizes ligands in the process of receptor-mediated endocytosis. This protein also acts as a co-receptor with Frizzled protein family members for transducing signals by Wnt proteins and was originally cloned on the basis of its association with type 1 diabetes mellitus in humans. This protein plays a key role in skeletal homeostasis and many bone density related diseases are caused by mutations in this gene. Mutations in this gene also cause familial exudative vitreoretinopathy. Alternative splicing results in multiple transcript variants.

Specifications

Catalog Number	KC-4179
Cell Line Name	293T LRP5 Cell Line
Host Cell Line	293T
Description	Stable 293T clone expressing exogenous human LRP5 gene in high level
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 LRP5 cell line was generated using a lentiviral vector expressing the human-LRP5 sequence.

Characterization

Figure 1: Characterization of human LRP5 overexpression in the 293T LRP5 stable clone using FACS.

Figure 2: Characterization of human LRP5 in the 293T LRP5 stable clone using PCR sequencing.

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

Ren Q, Chen J, Liu Y. LRP5 and LRP6 in Wnt Signaling: Similarity and Divergence. Front Cell Dev Biol. 2021 May 6;9:670960. doi: 10.3389/fcell.2021.670960. PMID: 34026761; PMCID: PMC8134664.
Nie X, Wang H, Wei X, Li L, Xue T, Fan L, Ma H, Xia Y, Wang YD, Chen WD. LRP5 Promotes Gastric Cancer via Activating Canonical Wnt/β-Catenin and Glycolysis Pathways. Am J Pathol. 2022 Mar;192(3):503-517. doi: 10.1016/j.ajpath.2021.11.007. Epub 2021 Dec 8. PMID: 34896072.