KC-0249

293T-human-CD47-Cell-Line

46437

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Background of 293T-human-CD47-Cell-Line

CD47, also known as integrin-associated protein (IAP), is a transmembrane protein that belongs to the immunoglobulin superfamily. It functions as a “don’t eat me” signal to macrophages by binding to its ligand, signal regulatory protein alpha (SIRPα). CD47 is considered a potential therapeutic target in certain cancers and in the treatment of pulmonary fibrosis.

Specifications

Catalog Number	KC-0249
Cell Line Name	293T-human-CD47-Cell-Line
Host Cell Line	293T
Description	Stable HEK293T clone expressing exogenous human CD47 gene
Quantity	One vial of frozen cells (≥2-10⁶/vial)
Stability	Stable in culture over a minimum of 10 passages
Application	Drug screening and biological assays
Freezing Medium	70% DMEM + 20% FBS + 10% DMSO
Propagation Medium	DMEM + 10% FBS + 0.5μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Epithelial
Subculture	Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL
Incubation	37 °C with 5% CO₂
Storage	Liquid nitrogen immediately upon receiving
Doubling Time	Approximately 30 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

293T human-CD47 cell line was generated using a lentiviral vector expressing the human-CD47 sequence.

Characterization

Figure 1: Characterization of human-CD47 overexpression in the 293T human-CD47 stable clone using FACS.

Cell Resuscitation

Prewarm culture medium (DMEM supplemented with 10% FBS and 0.5μg/mL puromycin)in a 37°C water bath.
Thaw the frozen vial in a 37°C water bath for 1-2 minutes.
Transfer the vial into biosafety cabinet, and wipe the surface with 70% ethanol.
Unscrew the top of the vial and transfer the cell suspension gently into a sterile centrifuge tube containing 9.0mL complete culture medium.
Spin at ~ 125 × g for 5-7 minutes at room temperature, and discard the supernatant without disturbing the pellet.
Resuspend cell pellet with the appropriate volume of complete medium and transfer the cell suspension into a T25 culture flask.
Incubate the flask at 37°C, 5% CO₂ incubator.
Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% DMEM + 20% FBS + 10% DMSO) fresh immediately before use.
Keep the freezing medium on ice and label cryovials.
Transfer cells to a sterile, conical centrifuge tube, and count the cells.
Centrifuge the cells at 250×g for 5 minutes at room temperature and carefully aspirate off the medium.
Resuspend the cells at a density of at least 3×10⁶ cells/mL in chilled freezing medium.
Aliquot 1 mL of the cell suspension into each cryovial.
Freeze cells in the CoolCell freezing container overnight in a -80°C freezer.
Transfer vials to liquid nitrogen for long-term storage.

References

Willingham, Stephen B, Jens-Peter Volkmer,et al. 2012.“The CD47-Signal Regulatory Protein Alpha (SIRPa) Interaction Is a Therapeutic Target for Human Solid Tumors.” Proceedings of the National Academy of Sciences of the United States of America 109(17). National Acad Sciences: 6662–67. doi:10.1073/pnas.1121623109.
Vonderheide, Robert H. 2015. “CD47 Blockade as Another Immune Checkpoint Therapy for Cancer.” Nature Medicine 21(10). Nature Publishing Group: 1122–23. doi:10.1038/nm.3965..

Use License Agreement

Research Use Only.

Not for use in diagnostic procedures or therapeutic applications.

Redistribution of the cell line or its derivatives is prohibited without prior written permission from Kyinno Biotechnology.