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Tumor cells can evade recognition and elimination by the body's immune system by utilizing immune checkpoint receptors; consequently, blocking these receptors holds promise as a broadly effective strategy for tumor immunotherapy. Although anti-PD-1/PD-L1 antibodies—much like anti-CTLA-4 antibodies—represent a relatively mature therapeutic modality, their overall efficacy in patients remains limited due to the prevalence of drug resistance. Therefore, the identification of novel targets for tumor immunotherapy has become a matter of urgent necessity.

Programmed Cell Death Protein 1 (PD-1) is a receptor expressed on activated T cells that binds to its ligands, PD-L1 and PD-L2, to negatively regulate immune responses. PD-1 ligands are present in the majority of cancers; the interaction between PD-1 and its ligands (PD-L1/2) suppresses T-cell activity and enables cancer cells to evade immune surveillance. The PD-1/PD-L1 signaling pathway constitutes a critical component of tumor-induced immunosuppression, serving to dampen T-lymphocyte activation and enhance immune tolerance toward tumor cells, thereby facilitating tumor immune evasion. The binding of PD-1 to PD-L1 can attenuate T-cell-mediated immune surveillance, leading to a compromised immune response or even inducing T-cell apoptosis. PD-1/PD-L1 inhibitors function by releasing anti-tumor T cells from this state of immunosuppression, thereby promoting T-cell proliferation, infiltration into the tumor microenvironment, and the subsequent induction of anti-tumor responses. Furthermore, the PD-1/PD-L1/2 pathway plays a role in regulating autoimmune responses, rendering these proteins promising therapeutic targets for a wide range of conditions, including various cancers as well as autoimmune diseases such as multiple sclerosis, arthritis, lupus, and type 1 diabetes.

The Jurkat E6.1 Human PD1/SHP1 Reporter cell Model—effectively simulates the signal transduction process of PD1/SHP1 *in vivo*. The underlying principle is illustrated in the figure below.

Figure 1. Schematic Diagram of the Jurkat E6.1 Human PD1/SHP1 Reporter cell Model

Jurkat E6.1 cell line expressing full length human PD1. Expression is confirmed by flow cytometry.

Figure 2. Recombinant PD-1/SHP1 reporter cell constitutively expressing PD1.

Figure 3. Blocking of PD1 induced PD1 SHP1 Reporter Cell (C1) Activity by PD1 Neutralizing/Atezolizumab with PD-L1 Target cell(Suspension)(C4).

Cell Passage Procedures

1.This cell line grows in suspension.
2.Upon receipt, cells should be thawed immediately or stored in liquid nitrogen until use.
3.Before thawing, pre-warm the water bath and culture medium to 37 °C, and prepare a small amount of dry ice.
4.Remove the cryovial from storage and transport it to the cell culture laboratory on dry ice.
5.Rapidly thaw the cells in a 37 °C water bath. Once the cells are completely thawed, spray the cryovial with 70% ethanol for disinfection and transfer it to a biosafety cabinet.
6.Add 10 mL of pre-warmed culture medium into a 15 mL centrifuge tube. Transfer the contents of the cryovial into the tube and centrifuge at 1000 rpm for 5 minutes.
7.Carefully discard the supernatant. Resuspend the cell pellet in 5 mL of pre-warmed culture medium by gentle pipetting. Immediately perform cell counting and adjust the cell density to 3–6 × 10⁵ cells/mL based on the counting results, then transfer the cells into a culture flask.
8.Count the cells every 1–2 days. When the cell density exceeds 1 × 10⁶ cells/mL, passage the cells promptly or add fresh culture medium. Maintain the cell density between 2 × 10⁵ and 1 × 10⁶ cells/mL.

Suspension Cell Cryopreservation Procedure:

1.Collect 8 × 10⁶ cells, centrifuge, and discard the supernatant.
2.Add 1 mL of cell freezing medium (90% FBS + 10% DMSO) and gently pipette to mix thoroughly. Transfer the suspension into a cryovial.
3.Immediately place the cryovial into a controlled-rate freezing container (Nalgene 5100-0001), fill with isopropanol up to the indicated level, and store at −80 °C.
4.After 24 hours, transfer the cryovial to liquid nitrogen for long-term storage.