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Receptor tyrosine kinases play a crucial role in normal cellular function. Humans have 58 known receptor tyrosine kinases, and one of the most important members of this family is c-KIT. c-KIT, also known as CD117, is a type III receptor tyrosine kinase encoded by the KIT gene. It uses stem cell growth factor (SCF) as a ligand and is therefore called the stem cell growth factor receptor (SCFR). c-KIT signaling is activated by binding to its ligand, the SCF protein, through autophosphorylation, which induces c-KIT dimerization and activation, initiating a phosphorylation cascade that regulates cell growth. c-KIT phosphorylation can trigger multiple signaling pathways, including the JAK/STAT, RAS/MAP kinase pathway, PI3 kinase, PLCγ pathway, and SRC pathway. Activation of the SCF/c-KIT signaling pathway is commonly associated with cell proliferation, migration, and survival, thereby regulating key functions such as hematopoiesis, pigmentation, and spermatogenesis.

The SCF/c-KIT Effector Reporter Cell model effectively mimics the in vivo SCF/c-KIT signaling pathway; the mechanism is illustrated in the figure below.

Figure 1. Schematic diagram of the HEK293 Human SCF/c-KIT(GNNK-) Effector Reporter Cell model

Figure 2. Recombinant SCF/c-KIT(GNNK-) Effector Reporter Cell stably expressing c-KIT.


Figure 3. Dose Response of Recombinant Human SCF in SCF/c-KIT(GNNK-) Effector Reporter Cell(C18).


Figure 4. Inhibition of Human SCF-induced Reporter Activity by c-KIT Blocking Ab in SCF/c-KIT(GNNK-) Effector Reporter Cell(C18).

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  HEK293 Human SCF/c-KIT(GNNK-) Effector Reporter 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.