KC-1630

Jurkat-NFAT-Luc2-TIGIT-Cell-Line

22503

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Background of Jurkat-NFAT-Luc2-TIGIT-Cell-Line

T cell immunoglobulin and ITIM domain (TIGIT) is an inhibitory receptor expressed on lymphocytes that was recently propelled under the spotlight as a major emerging target in cancer immunotherapy. TIGIT interacts with CD155 expressed on antigen-presenting cells or tumour cells to down-regulate T cell and natural killer (NK) cell functions. Here we identify TIGIT as a coinhibitory receptor that critically limits antitumor and other CD8(+) T cell-dependent chronic immune responses. TIGIT is highly expressed on human and murine tumor-infiltrating T cells, and, in models of both cancer and chronic viral infection, antibody coblockade of TIGIT and PD-L1 synergistically and specifically enhanced CD8(+) T cell effector function, resulting in significant tumor and viral clearance, respectively.

Specifications

Catalog Number	KC-1630
Cell Line Name	Jurkat-NFAT-Luc2-TIGIT-Cell-Line
Host Cell Line	Jurkat
Description	Stable Jurkat cell line expressing exogenous luciferase gene under the control of TIGIT signaling pathway
Quantity	Two vials of frozen cells (≥2-10⁶/vial)
Stability	Stable in culture over a minimum of 10 passages
Application	Cell model for monitoring EGFR signaling pathway
Freezing Medium	70% RPMI1640+20% FBS+10% DMSO
Propagation Medium	RPMI1640+10%FBS+300µg/mL Hygromycin B+0.75µg/mL Puromycin
Selection Marker	Puromycin, HygromycinB
Morphology	Lymphoblast
Subculture	Split saturated culture 1:4-1:6 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 26 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

Jurkat-NFAT-Luc2-TIGIT cell line was generated using lentivirus expressing TIGIT sequence.

Characterization

Figure 1. Characterization of TIGIT over-expression in Jurkat-NFAT-Luc2 stable clones using FCAS.

Figure 2. Jurkat-NFAT-Luc2-TIGIT cell line was seeded into the 96-well plate, and co-culture with 293T-OS8-PDL1 for 6 hours, then readout with Bright-Glo system.

Figure 3.Jurkat-NFAT-Luc2-TIGIT cell line was seeded into the 96-well plate, and treated with KA-1177-Tiragolumab, KA-1016-Atezolizumab and KP2001-hIgG1 isotype at a maximum concentration 4 μg/mL for 1 hours, and co-culture with 293T-OS8-PDL1 for 6 hours, then readout with Bright-Glo system.

Cell Resuscitation

Prewarm culture medium (RPMI1640 + 10% FBS + 300µg/mL Hygromycin B + 0.75µg/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 1:4-1:6 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% RPMI1640 + 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

Harjunpää H, Guillerey C. TIGIT as an emerging immune checkpoint. Clin Exp Immunol. 2020 May;200(2):108-119.
Johnston RJ, Comps-Agrar L, Hackney J, Yu X, Huseni M, Yang Y, Park S, Javinal V, Chiu H, Irving B, Eaton DL, Grogan JL. The immunoreceptor TIGIT regulates antitumor and antiviral CD8(+) T cell effector function. Cancer Cell. 2014 Dec 8;26(6):923-937.