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SIRPα (Signal Regulatory Protein α) is an immunoglobulin superfamily transmembrane protein primarily expressed on the surface of myeloid cells (such as macrophages and dendritic cells). Its extracellular region contains three Ig-like domains (D1, D2, D3), with the D1 domain specifically binding to CD47. The intracellular segment includes an immunoreceptor tyrosine-based inhibitory motif (ITIM), which can recruit SHP-1/SHP-2 phosphatases to transmit inhibitory signals. SIRPα maintains self-immune tolerance by regulating phagocytic activity, preventing macrophages from mistakenly damaging normal cells.

  
CD47 is a five-transmembrane protein widely expressed on the surface of red blood cells, platelets, and tumor cells. Its extracellular N-terminal IgV-like domain binds to SIRPα and is known as the 'Don't eat me' signaling molecule, playing a key role in red blood cell homeostasis (preventing clearance of young red blood cells by splenic macrophages) and tumor immune escape.

  
When CD47 binds to SIRPα, the ITIM motif in SIRPα is phosphorylated, recruiting SHP-1/SHP-2 phosphatases. These phosphatases inhibit downstream pro-phagocytic signals through dephosphorylation.

The Jurkat E6.1 Human SIRPα&CD47 Dual Effector Reporter Cell model effectively mimics the in vivo SIRPα signaling transduction process. The principle is illustrated in the figure below.

Figure 1. Schematic diagram of the Jurkat E6.1 Human SIRPα&CD47 Dual Effector Reporter Cell model.

Figure 2. Dose Response of Blocking Antibodies in SIRPɑ&CD47 Dual Effector Reporter Cell(C33). Dose Response of CD47 Blocking Antibody in SIRPɑ&CD47 Dual Effector Reporter Cell(C33).

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.