KC-1008

293T-Cyno-CD122-Cell-Line

43804

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Background of 293T-Cyno-CD122-Cell-Line

"CD122, also named as Interleukin-2 receptor subunit beta (IL2RB), which is involved in T cell-mediated immune responses, is present in 3 forms with respect to ability to bind interleukin 2. The low affinity form is a monomer of the alpha subunit (also called CD25) and is not involved in signal transduction. The intermediate affinity form consists of a gamma/beta subunit heterodimer, while the high affinity form consists of an alpha/beta/gamma subunit heterotrimer. Both the intermediate and high affinity forms of the receptor are involved in receptor_x0002_mediated endocytosis and transduction of mitogenic signals from interleukin 2. The protein encoded by this gene represents the beta subunit and is a type I membrane protein. This protein also forms one of the three subunits of the IL-15 receptor. Activation of the receptor increases proliferation of CD8+ effector T cells."

Specifications

Catalog Number	KC-1008
Cell Line Name	293T-Cyno-CD122-Cell-Line
Host Cell Line	293T
Description	HEK293T cell line stably expressing exogenous cyno CD122 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 + 1 μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Fibroblastoid cells growing as monolayer
Subculture	Split saturated culture 1:4-1:5 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 CD122 cell line was generated using lentiviral vector expressing monkey CD122 sequence.

Characterization

Figure 1: Characterization of CD122 overexpressing in 293T stable clones using FACS.

Cell Resuscitation

Prewarm culture medium (DMEM supplemented with 10% FBS and 1 μg/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 1:4-1:5 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

Boyman O, Sprent J (February 17, 2012). The role of interleukin-2 during homeostasis and activation of the immune system. Nat Rev Immunol. 12 (3): 180–190.
Miyazaki T, Kawahara A, Fujii H, Nakagawa Y, Minami Y, Liu ZJ, Oishi I, Silvennoinen O, Witthuhn BA, Ihle JN (November 1994). Functional activation of Jak1 and Jak3 by selective association with IL-2 receptor subunits. Science. 266 (5187): 1045–7.