H_SIRPα Reporter Jurkat Cell Line

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Current position：Product Center > Cell lines > Immune checkpoints > CD47-SIRPα > H_SIRPα Reporter Jurkat Cell Line

H_SIRPα Reporter Jurkat Cell Line

Description

Cat. No: GM-C28270
Product: H_SIRPα Reporter Jurkat Cell Line

SIRPα (Signal Regulatory Protein Alpha) is a cell surface protein in the immunoglobulin superfamily, mainly expressed on immune cells like macrophages and dendritic cells. It regulates immune responses by binding to its ligand CD47, primarily inhibiting macrophage phagocytosis, which is essential for immune surveillance and self-tolerance.

In signaling pathways, SIRPα activates downstream mechanisms through CD47 binding, inhibiting Src family tyrosine kinases. This interaction leads to the phosphorylation of SIRPα's intracellular domain, reducing macrophage activation and phagocytosis. SIRPα also affects immune cell function and tumor microenvironment formation by regulating cytokine release and intercellular interactions, making it important for research on tumor immune evasion and autoimmune diseases.

H_SIRPα Reporter Jurkat Cell Line is a clonal stable Jurkat cell line constructed using lentiviral technology, constitutive expression of the human SIRPα, and exhibits signal-dependent expression of a luciferase reporter gene. The reporter cell line is co-cultured with the H_CD47 aAPC CHO-K1 Cell Line (Cat. GM-C13353). The interaction between CD47 and SIRPα inhibits TCR-CD3 signaling. By adding Anti-CD47 and Anti-SIRPα antibodies, the interactions of CD47-SIRPα are blocked, thereby restoring T cell signaling. The luciferase readout indicates the activation level of the signaling pathway, allowing evaluation of the in vitro effects of CD47-SIRPα related drugs.

Data

Response to Anti-CD47 hIgG4 Antibody(5F9). Serial dilutions of the Anti-CD47 hIgG4 Antibody(5F9) (Cat. GM-27657AB) were incubated with 1E4 cells/well of the H_CD47 aAPC CHO-K1 Cell Line (Cat. GM-C13353) in a 96-well plate for 1 hour in assay buffer (RPMI 1640 + 1% FBS + 1% P.S). Subsequently, the H_SIRPα Reporter Jurkat Cell Line (Cat. GM-C28270) at a concentration of 1E5 cells/well was added, and the co-culture proceeded for an additional 16 hours. Firefly luciferase activity is then measured using the GMOne-Step Luciferase Reporter Gene Assay Kit (Cat. GM-040503). The results indicated a maximum blocking fold of approximately [11.6]. Data are shown by drug mass concentration.

Response to Anti-H_SIRPα hIgG1 Antibody(Hu1H9-G1). H_CD47 aAPC CHO-K1 Cell Line (Cat. GM-C13353) was seeded at a density of 1E4 cells/well in a 96-well plate and incubated overnight. The next day, serial dilutions of the Anti-H_SIRPα hIgG1 Antibody(Hu1H9-G1) (Cat. GM-49522AB) were incubated with 1E5 cells/well of the H_SIRPα Reporter Jurkat Cell Line (Cat. GM-C28270) in a 96-well plate for 1 hour, and then added to the pre-seeded cells. The mixture was incubated for an additional 16 hours. Firefly luciferase activity is then measured using the GMOne-Step Luciferase Reporter Gene Assay Kit (Cat. GM-040503). The results indicated maximum folds of approximately [20.6]. Data are shown by drug mass concentration.

The Sanger sequencing of the H_SIRPα Reporter Jurkat Cell Line showed successful knockout of CD47

H_SIRPα Reporter Jurkat Cell Line (H_CD47 KO) was determined by flow cytometry using Anti-CD47 hIgG4 Antibody(5F9) (Cat. GM-27657AB).

Specifications

Cat. No		GM-C28270
Product		H_SIRPα Reporter Jurkat Cell Line
Product Format		1 vial of frozen cells
Quantity		5E6 Cells per vial,1 mL
Storage Conditions		Liquid nitrogen immediately upon receipt
Recovery Medium		RPMI 1640+10% FBS+1% P.S
Growth medium		RPMI 1640+10% FBS+1% P.S+3.5 μg/mL Blasticidin+400 μg/mL Bleomycin+200 μg/mL Hygromycin+0.75 μg/mL Puromycin
Note		None
Freezing Medium		90% FBS+10%DMSO
Growth properties		Suspension
Growth Conditions		37°C, 5% CO₂
Safety considerations		Biosafety Level 2
Mycoplasma Testing		The cell line has been screened to confirm the absence of Mycoplasma species.

Materials

Reagent		Ordering Information
RPMI 1640		VivaCell/C3010-0500
Fetal Bovine Serum		Cegrogen biotech/A0500-3010
Pen/Strep		Thermo/15140-122
Blasticidin		Genomeditech/GM-040404
Bleomycin		Genomeditech/GM-040407
Hygromycin		Genomeditech/GM-040403
Puromycin		Genomeditech/GM-040401
H_CD47 aAPC CHO-K1 Cell Line		Genomeditech/GM-C13353
Anti-CD47 hIgG4 Antibody(5F9)		Genomeditech/GM-27657AB
Anti-H_SIRPα hIgG1 Antibody(Hu1H9-G1)		Genomeditech/GM-49522AB
GMOne-Step Luciferase Reporter Gene Assay Kit		Genomeditech/GM-040503

Cell Culture

Cell Recovery

Recovery Medium: RPMI 1640+10% FBS+1% P.S

To insure the highest level of viability, thaw the vial and initiate the culture as soon as possible upon receipt. If upon arrival, continued storage of the frozen culture is necessary, it should be stored in liquid nitrogen vapor phase and not at -70°C. Storage at -70°C will result in loss of viability.

a) Thaw the vial by gentle agitation in a 37°C water bath. To reduce the possibility of contamination, keep the O-ring and cap out of the water. Thawing should be rapid (approximately 2 - 3 minutes).

b) Remove the vial from the water bath as soon as the contents are thawed, and decontaminate by dipping in or spraying with 70% ethanol. All of the operations from this point on should be carried out under strict aseptic conditions.

c) Transfer the vial contents to a centrifuge tube containing 5.0 mL complete culture medium. And spin at approximately 176 x g for 5 minutes. Discard supernatant.

d) Resuspend cell pellet with the recommended complete medium. And dispense the suspension into 1 - 2 T-25 culture flasks.

e) Incubate the culture at 37°C in a suitable incubator. A 5% CO₂ in air atmosphere is recommended if using the medium described on this product sheet.

Cell Freezing

Freezing Medium: 90% FBS+10%DMSO

a) Centrifuge at 176 x g for 3 minutes to collect cells.

b) Resuspend the cells in pre-cooled freezing medium and adjust the cell density to 5E6 cells/mL.

c) Aliquot 1 mL into each vial.

d) Place the vial in a controlled-rate freezing container and store at -80°C for at least 1 day, then transfer to liquid nitrogen as soon as possible.

Cell passage

Growth medium: RPMI 1640+10% FBS+1% P.S+3.5 μg/mL Blasticidin+400 μg/mL Bleomycin+200 μg/mL Hygromycin+0.75 μg/mL Puromycin

Approximately 48-72 hours after the initial thawing, the cells can be passaged for the first time. After this initial passage, the culture medium can be adjusted to growth medium supplemented with antibiotics. If cells are not passaged within 48 hours, it is recommended to add some fresh recovery medium and place the flask horizontally.

a) When the cell density reaches 1.5 - 2E6 cells/mL, subculture the cells. Do not allow the cell density to exceed 2E6 cells/mL.

b) It is recommended to use T-25 flasks for subculturing.

c) These cells are suspension cells, and it is recommended to use the "half-medium change" method to maintain optimal cell conditions during passaging.

d) During passaging, you can directly add fresh growth medium to the culture flask, gently pipette to resuspend the cells, and then transfer the cell suspension to a new T-25 flask for continued culture.

Subcultivation Ratio: Maintain cultures at a cell concentraion between 3E5 and 1E6 viable cells/mL.

Medium Renewal: Every 2 to 3 days

Notes

a) These cells are sensitive to density, so please ensure that the cell density is maintained within an appropriate range during culture and subculturing.

b) During the first passage, pay attention to the nutrient supply; if not subculturing, make sure to add fresh recovery medium every other day as needed.

Cell Recovery

Recovery Medium: RPMI 1640+10% FBS+1% P.S

c) Transfer the vial contents to a centrifuge tube containing 5.0 mL complete culture medium. And spin at approximately 176 x g for 5 minutes. Discard supernatant.

d) Resuspend cell pellet with the recommended complete medium. And dispense the suspension into 1 - 2 T-25 culture flasks.

e) Incubate the culture at 37°C in a suitable incubator. A 5% CO₂ in air atmosphere is recommended if using the medium described on this product sheet.

Cell Freezing

Freezing Medium: 90% FBS+10%DMSO

a) Centrifuge at 176 x g for 3 minutes to collect cells.

b) Resuspend the cells in pre-cooled freezing medium and adjust the cell density to 5E6 cells/mL.

c) Aliquot 1 mL into each vial.

d) Place the vial in a controlled-rate freezing container and store at -80°C for at least 1 day, then transfer to liquid nitrogen as soon as possible.

Cell passage

Growth medium: RPMI 1640+10% FBS+1% P.S+3.5 μg/mL Blasticidin+400 μg/mL Bleomycin+200 μg/mL Hygromycin+0.75 μg/mL Puromycin

a) When the cell density reaches 1.5 - 2E6 cells/mL, subculture the cells. Do not allow the cell density to exceed 2E6 cells/mL.

b) It is recommended to use T-25 flasks for subculturing.

c) These cells are suspension cells, and it is recommended to use the "half-medium change" method to maintain optimal cell conditions during passaging.

Subcultivation Ratio: Maintain cultures at a cell concentraion between 3E5 and 1E6 viable cells/mL.

Medium Renewal: Every 2 to 3 days

Notes

a) These cells are sensitive to density, so please ensure that the cell density is maintained within an appropriate range during culture and subculturing.

b) During the first passage, pay attention to the nutrient supply; if not subculturing, make sure to add fresh recovery medium every other day as needed.

CD47:SIRPα
	H_SIRPα Blockade Reporter Cell Line	Cynomolgus_CD47 CHO-K1 Cell Line
H_CD47 aAPC CHO-K1 Cell Line	H_CD47 CHO-K1 cell line
H_CD47 MC38 Cell Line	H_CD47 PDL1 MC38(mouse_PDL1 KO) Cell Line	H_SIRPA(SIRPα) CHO-K1 Cell Line
Mouse_CD47 CHO-K1 Cell Line
Anti-CD47 hIgG4 Antibody(5F9)

CD47:SIRPα
	H_SIRPα Blockade Reporter Cell Line	Cynomolgus_CD47 CHO-K1 Cell Line
H_CD47 aAPC CHO-K1 Cell Line	H_CD47 CHO-K1 cell line
H_CD47 MC38 Cell Line	H_CD47 PDL1 MC38(mouse_PDL1 KO) Cell Line	H_SIRPA(SIRPα) CHO-K1 Cell Line
Mouse_CD47 CHO-K1 Cell Line
Anti-CD47 hIgG4 Antibody(5F9)