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SIRPα (Signal Regulatory Protein α) is a transmembrane protein belonging to the immunoglobulin superfamily. It is primarily expressed on the surface of myeloid cells (such as macrophages and dendritic cells). Its extracellular region contains three immunoglobulin-like domains (D1, D2, and D3), of which the D1 domain specifically binds to CD47. Its intracellular segment contains Immunoreceptor Tyrosine-based Inhibitory Motifs (ITIMs), which recruit SHP-1 and SHP-2 phosphatases to transmit inhibitory signals. By regulating phagocytic activity, SIRPα maintains self-tolerance and prevents macrophages from inadvertently attacking normal cells.    

CD47 is a five-pass transmembrane protein widely expressed on the surface of erythrocytes, platelets, and tumor cells. Its extracellular N-terminal IgV-like domain binds to SIRPα; consequently, it is known as a "Don’t eat me" signaling molecule. It plays a critical role in erythrocyte homeostasis (specifically, preventing splenic macrophages from clearing young erythrocytes) and in tumor immune evasion. 

Upon binding to CD47, the intracellular ITIM motifs of SIRPα undergo phosphorylation, thereby recruiting SHP-1 and SHP-2 phosphatases to inhibit downstream pro-phagocytic signals through dephosphorylation.

The SIRPα Effector Reporter Cell model accurately simulates the in vivo signal transduction process of SIRPα, as illustrated in the figure below.    

Figure 1. Schematic Diagram of the SIRPα Effector Reporter Cell Model

Figure 2. Dose response of SIRPɑ Blocking Antibodies in SIRPɑ Effector Reporter Cell(C11) With CD47⁺ Target Cell. Dose response of CD47 Blocking Antibody in SIRPɑ Effector Reporter Cell(C11) With CD47⁺ Target Cell.

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.