KC-2150

A549-CD228-Cell-Line

23486

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Background of A549-CD228-Cell-Line

CD228, also named as MFI2, is a member of the transferrin superfamily that binds to a single ferric iron with high affinity. CD228 expression is low in normal tissues but high in tumor and embryonic tissue. Soluble CD228 (sMTF) has also been reported to be increased in patients with AD or arthritis.

Specifications

Catalog Number	KC-2150
Cell Line Name	A549-CD228-Cell-Line
Host Cell Line	Human A549 cell line
Description	A549 cell line stable expressing exogenous CD228 gene
Quantity	Two vials 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% McCoy's 5a+20% FBS+10% DMSO
Propagation Medium	McCoy5A+10%FBS+1μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Epithelial
Subculture	Split saturated culture 1:3-1:8 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 27 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

A549 human CD228 cell line was generated using a lentiviral vector expressing human CD228 sequence.

Characterization

Cell Resuscitation

Prewarm culture medium (Mycoy5A + 10% FBS + 1μg/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 1:3-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% McCoy's 5a + 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

Suryo Rahmanto, Y., Dunn, L. L. & Richardson, D. R. The melanoma tumor antigen, melanotransferrin (p97): a 25-year hallmark ÿ from iron metabolism to tumorigenesis. Oncogene 26, 6113ÿ6124 (2007).
SAWAKI, K. et al. Level of Melanotransferrin in Tissue and Sera Serves as a Prognostic Marker of Gastric Cancer. Anticancer Res. 39, 6125ÿ6133 (2019).