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Tumor Necrosis Factor-alpha (TNF-α) is a transmembrane protein with a molecular weight of 26 kDa; upon cleavage of its precursor peptide by TNF-α-converting enzyme (TACE), it is secreted as a soluble 17 kDa molecule. Although activated macrophages are the primary source of TNF-α, it can also be produced by a variety of other cell types, such as fibroblasts, astrocytes, smooth muscle cells, keratinocytes, and various tumor cells—including B-cell lymphomas, breast cancer, and colon cancer cells.

Acting as a tumor-promoting factor, TNF-α participates in every stage of tumorigenesis, including cellular transformation, proliferation, angiogenesis, invasion, and metastasis. TNF-α signaling also constitutes a critical component of the immune system, capable of suppressing tumorigenesis and inhibiting viral replication; furthermore, it functions as an endogenous pyrogen, capable of inducing fever and apoptosis.

Membrane-Anchored TNF-α Cells (Adherent): Stably express human TNF-α.

Figure 1. Recombinant Membrane Anchored TNFα Cell (Adherent) constitutively expressing TNFα.

Figure 2. Dose Response of TNFα Abs in ADCC Bioassay Effector Cell V variant (High Affinity)-Fcγ-NFAT-Jurkat (C36) with Membrane Anchored TNFα Cell (Adherent, C12).

Cell Resuscitation
1)Rapidly thaw the frozen cells in a 37 °C water bath for approximately 60 seconds. Once thawed (which may take slightly less or more than 60 seconds), immediately transfer the cell suspension from the cryovial into a 15 mL centrifuge tube containing 10 mL of pre-warmed  CHO-K1 Human Membrane Anchored TNFα Cell (Adherent) Cell complete culture medium.
2)Centrifuge cells at 1000 rpm for 5 min to remove medium, then resuspend cells in 5 mL of pre-warmed complete medium.
3)Transfer the cell suspension into a T25 culture flask and incubate at 37 °C with 5% CO₂.
4)After approximately 24–36 hours, replace the medium or passage the cells to remove non-adherent dead cells.

Subculturing procedure
1)When the cell density reaches the appropriate confluency for passaging, wash the cells with PBS, then add 1 mL trypsin to detach the cells. When more than 80% of the cells detach upon gently tapping the culture flask, add complete culture medium to terminate digestion. Gently pipette to obtain a single-cell suspension, transfer to a 15 mL centrifuge tube, and centrifuge at 1000 rpm for 5 minutes.

2)Discard supernatant after centrifugation. Resuspend cells in fresh medium to a single-cell suspension and transfer to a new culture flask for continued growth.

Cell Freezing
After trypsinization and centrifugation of cells from each T75 flask or 10 cm culture dish, discard the supernatant. Add 2 mL of cryopreservation medium (90% FBS + 10% DMSO), gently resuspend thoroughly, and aliquot into two cryovials. Immediately place the cryovials into a controlled-rate freezing container (e.g., Nalgene 5100-0001), fill with isopropanol to the indicated level, and store at −80 °C. After 24 hours, transfer the cryovials to liquid nitrogen for long-term storage.