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TIGIT—also known as WUCAM, Vstm3, or VSIG9—is a co-inhibitory receptor belonging to the immunoglobulin superfamily. High levels of TIGIT expression are observed on effector CD4+ and CD8+ T cells, regulatory T cells, and NK cells. TIGIT's ligands include CD112 and the Poliovirus Receptor (PVR—also known as CD155, Necl-5, and Tage4), with PVR serving as TIGIT's high-affinity cognate receptor. Furthermore, TIGIT competes with CD226 (DNAM-1) and CD96 (TACTILE) for their respective ligands. TIGIT effectively blocks the binding of CD155 to either CD96 or CD226, thereby further substantiating that TIGIT possesses the highest affinity for CD155. TIGIT not only competes with CD226 for ligands but can also directly bind to CD226 *in cis*, thereby preventing its homodimerization and rendering CD226 unable to bind to CD155 to exert its co-stimulatory function. 

PVRIG (CD112R) is a co-inhibitory receptor within the Poliovirus Receptor (PVR) family, expressed on natural killer (NK) cells and T cells. It binds to its ligand (CD112 or PVRL2/nectin-2) to suppress T-cell and NK-cell responses against cancer. Additionally, PVRIG exerts immunosuppressive effects by competitively binding to the ligands of CD226, thereby blocking the stimulatory signals transmitted by CD226. This mechanism of immunosuppression is similar to that employed by TIGIT—another member of the same receptor family—and it is possible that the two receptors act synergistically to mediate immunosuppression. Consequently, CD112R holds significant potential as a novel immune checkpoint inhibitor in the field of cancer immunotherapy. Serving as target cells for the TIGIT/PVRIG Dual Effector Reporter Cell, the CD155/CD112 Dual aAPC Cells effectively simulate the in vivo signal transduction processes of TIGIT and PVRIG. The underlying principle is illustrated in the figure below.

Figure 1. Schematic Diagram of the CD155/CD112 Dual aAPC Cell Model

Figure 2. Dose Response of Blocking Antibodies in TIGIT/PVRIG Dual Effector Reporter Cell(C45) with CD112/CD155 Dual aAPC Cell.

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  CHO-K1 Human CD155&CD112 Dual aAPC 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.