L-WRN Cells

3D cell culture

The L-WRN cell line is a murine fibroblast cell line derived from the L cells, which are mouse fibroblasts originally isolated from connective tissue. L-WRN cells have been engineered to stably express Wnt3a, R-spondin 3, and Noggin. These factors are critical for the growth and maintenance of intestinal organoids and stem cell cultures. The overexpression of these proteins enhances the proliferation and differentiation of intestinal stem cells, making L-WRN cells a valuable tool for studying intestinal biology and disease modeling.

In addition to their application in organoid culture, L-WRN cells serve as a robust model for investigating Wnt signaling pathways. Wnt signaling is pivotal in regulating cell fate, proliferation, and migration during development and in adult tissues. By providing a consistent and controlled source of Wnt3a, R-spondin 3, and Noggin, L-WRN cells facilitate research into the molecular mechanisms underlying these processes. Researchers can use these cells to dissect the roles of these signaling molecules in various biological contexts, including cancer, tissue regeneration, and developmental biology.

Overall, the L-WRN cell line is a powerful tool in biomedical research due to its ability to support the growth of complex three-dimensional cultures and its utility in studying key signaling pathways. Its role in the advancement of intestinal stem cell research and its contributions to our understanding of Wnt signaling highlight its importance in the field of cellular and molecular biology.

Adherent

Male

Fibroblast

DMEM, w: 4.5 g/L Glucose, w: 4 mM L-Glutamine, w: 1.5 g/L NaHCO3, w: 1.0 mM Sodium pyruvate (Cytion article number 820300a)

Remove the old medium from the adherent cells and wash them with PBS that lacks calcium and magnesium. For T25 flasks, use 3-5 ml of PBS, and for T75 flasks, use 5-10 ml. Then, cover the cells completely with Accutase, using 1-2 ml for T25 flasks and 2.5 ml for T75 flasks. Let the cells incubate at room temperature for 8-10 minutes to detach them. After incubation, gently mix the cells with 10 ml of medium to resuspend them, then centrifuge at 300xg for 3 minutes. Discard the supernatant, resuspend the cells in fresh medium, and transfer them into new flasks that already contain fresh medium.

Confirm that the vial remains deeply frozen upon delivery, as cells are shipped on dry ice to maintain optimal temperatures during transit.

Upon receipt, either store the cryovial immediately at temperatures below -150° C to ensure the preservation of cellular integrity, or proceed to step 3 if immediate culturing is required.

For immediate culturing, swiftly thaw the vial by immersing it in a 37° C water bath with clean water and an antimicrobial agent, agitating gently for 40-60 seconds until a small ice clump remains.

Perform all subsequent steps under sterile conditions in a flow hood, disinfecting the cryovial with 70% ethanol before opening.

Carefully open the disinfected vial and transfer the cell suspension into a 15 ml centrifuge tube containing 8 ml of room-temperature culture medium, mixing gently.

Centrifuge the mixture at 300 x g for 3 minutes to separate the cells and carefully discard the supernatant containing residual freezing medium.

Gently resuspend the cell pellet in 10 ml of fresh culture medium. For adherent cells, divide the suspension between two T25 culture flasks; for suspension cultures, transfer all the medium into one T25 flask to promote effective cell interaction and growth.

Adhere to established subculture protocols for continued growth and maintenance of the cell line, ensuring reliable experimental outcomes.

When planning to store a cryovial in liquid nitrogen for future thawing, it is mandatory to adhere to stringent safety measures. Appropriate protective gloves and clothing are essential, and the use of a face mask or safety goggles is required during the transfer of frozen samples to or from the liquid nitrogen tank. This is to mitigate the risk of injury from potential cryovial explosions upon removal, which can result in the projection of sharp fragments.

We stand by the promise of delivering products with high cell viability and robust culture performance. To achieve the best results, please make sure you follow the storage and culture instructions detailed in the product information sheet closely. Your adherence to these guidelines is key to success.