IL-2R is made up of three chains, α (CD25), β (CD122), and γ (CD132), of which only the α-chain is specific for IL-2. Signaling takes place mainly via CD122. Activated T cells express the high affinity receptor composed of all three chains. However, cells other than T cells, such as monocytes, express functional CD122 and CD132 and have the ability to signal following ligation of the intermediate affinity receptor consisting of the heterodimer formed by CD122 and CD132.
Ba/F3 cell, a murine interleukin-3 dependent pro-B cell line, is a popular system for exploring both kinases and their inhibitors, because some protein kinases can render the Ba/F3 cells to be depended on the activation of the kinases instead of IL-3 supplement, while their inhibitors can antagonize the kinase-dependent growth effects.
Specifications
Catalog Number:
KC-2450
Cell Line Name:
Ba/F3-STAT5-Luc2-IL2RG-reporter Cell Line
Host Cell Line:
Mouse Ba/F3 cell line
Description:
Stable Ba/F3 cell line expressing exogenous human IL2RG gene and luciferase under the control of STAT5 signaling pathway.
Mostly single, round (some polymorph) cells in suspension
Subculture:
Split saturated culture 1:10 every 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 20 hours
Mycoplasma Status:
Negative
Cell Line Generation
Ba/F3-STAT5-Luc2-IL2RG Cell Line was generated using a lentiviral vector expressing the human IL2RG sequence.
Characterization
Application
Cell Resuscitation
1. Prewarm culture medium (RPMI1640 + 10% FBS + 1μg/mL Puromycin + 1mg/mL Hygromycin B + 8ng/mL mouse IL-3) 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 1:10 every 3 days; seed out at about 1-3 × 105 cells/mL.
Cell Freezing
1. Prepare the freezing medium (70% RPMI-1640 + 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. James W. Verbsky, John R. Routes, Infections, Immune Disorders, and Autoinflammatory Diseases. Nelson Pediatric Symptom-Based Diagnosis, 2018.
