KC-1649

293T-IL2RB-IL2RG Cell Line

22541

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Background of 293T-IL2RB-IL2RG Cell Line

Interleukin-2 (IL-2) has emerged as the quintessential immunoregulatory cytokine with dual roles in both promotion of host defense and immune tolerance. The interleukin 2 receptor, which is involved in T cell-mediated immune responses, is present in 3 forms with respect to the ability to bind interleukin 2. The high-affinity IL-2R consists of three polypeptide chains: alpha (IL-2Rα, CD25), beta (IL-2Rβ, CD122), and gamma (IL-2Rγ, γc, CD132). IL-2Rβ and IL2Rγare required for downstream signal transduction by IL-2, and when expressed in the absence of IL-2Rα form the intermediate-affinity IL-2R which binds to IL-2 with ~100-fold lower affinity than the high affinity trimeric receptor. immune cells, like monocytes and B-cells, and nonimmune cells, like endothelial cells, may express different compositions of the IL-2R complex. A variety of drugs and triple antibodies against this target are in clinical stage.

Specifications

Catalog Number	KC-1649
Cell Line Name	293T-IL2RB-IL2RG Cell Line
NCBI/UniProt Accession Number	NM_000878.5 & NM_000206.3
Clone Number	1#
Host Cell Line	Human HEK293T cell line
Description	Stable HEK293T clone expressing exogenous human IL2RB-IL2RG
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-IL2RB-IL2RG cell line was generated using a lentiviral vector expressing the human IL2RB and IL2RG sequence.

Characterization

Figure: Characterization of human IL2RB overexpression in 293T stable clones using FACS.

Cell Resuscitation

1. Prewarm culture medium (DMEM supplemented with 10% FBS and 1μg/mL Puromycin)in a 37°C water bath.
2. Thaw the frozen vial in a 37°C water bath for 1-2 minutes.
3. Transfer the vial into biosafety cabinet, and wipe the surface with 70% ethanol.
4. Unscrew the top of the vial and transfer the cell suspension gently into a sterile centrifuge tube containing 9.0mL complete culture medium.
5. Spin at ~ 125 × g for 5-7 minutes at room temperature, and discard the supernatant without disturbing the pellet.
6. Resuspend cell pellet with the appropriate volume of complete medium and transfer the cell suspension into a T25 culture flask.
7. Incubate the flask at 37°C, 5% CO₂ incubator.
8. Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

1. Prepare the freezing medium (70% DMEM + 20% FBS + 10% DMSO) fresh immediately before use.
2. Keep the freezing medium on ice and label cryovials.
3. Transfer cells to a sterile, conical centrifuge tube, and count the cells.
4. Centrifuge the cells at 250×g for 5 minutes at room temperature and carefully aspirate off the medium.
5. Resuspend the cells at a density of at least 3×10⁶ cells/mL in chilled freezing medium.
6. Aliquot 1 mL of the cell suspension into each cryovial.
7. Freeze cells in the CoolCell freezing container overnight in a -80°C freezer.
8. Transfer vials to liquid nitrogen for long-term storage

References

1. Damoiseaux J. The IL-2 - IL-2 receptor pathway in health and disease: The role of the soluble IL-2 receptor. Clin Immunol. 2020 Sep;218:108515. doi: 10.1016/j.clim.2020.108515. Epub 2020 Jul 1. PMID: 32619646.
2. Hernandez JD, Hsieh EWY. A great disturbance in the force: IL-2 receptor defects disrupt immune homeostasis. Curr Opin Pediatr. 2022 Dec 1;34(6):580-588. doi: 10.1097/MOP.0000000000001181. Epub 2022 Sep 27. PMID: 36165614; PMCID: PMC9633542.