KC-5967

293T-chimeric-STEAP1-loop1-Flag Cell Line

60639

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Background of 293T-chimeric-STEAP1-loop1-Flag Cell Line

This gene is predominantly expressed in prostate tissue, and is found to be upregulated in multiple cancer cell lines. The gene product is predicted to be a six-transmembrane protein, and was shown to be a cell surface antigen significantly expressed at cell-cell junctions.

Specifications

Catalog Number	KC-5967
Cell Line Name	293T-chimeric-STEAP1-loop1-Flag Cell Line
NCBI/UniProt Accession Number	NM_012449.3
Clone Number	12#
Host Cell Line	293T
Description	Stable 293T cell line expressing exogenous chimeric-STEAP1-loop1-Flag 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% DMEM+20% FBS+10% DMSO
Propagation Medium	DMEM+10% FBS +1μg/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

Cell Line Generation

293T-chimeric-STEAP1-loop1-Flag cell line was generated using lentivirus expressing chimeric-STEAP1-loop1-Flag sequence.

Characterization

Figure 1. Characterization of chimeric-STEAP1-loop1-Flag over-expression in the 293T-chimeric-STEAP1-loop1-Flag stable clone using FACS.

Figure 2.Characterization of 293T-chimeric-STEAP1-loop1-Flag cell line stable clone using PCR sequencing.

Cell Resuscitation

Prewarm culture medium (DMEM supplemented with 10% FBS and 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 1:4-1:8 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

Zhang Z, Hou WB, Zhang C, Tan YE, Zhang DD, An W, Pan SW, Wu WD, Chen QC, Xu HM. A research of STEAP1 regulated gastric cancer cell proliferation, migration and invasion in vitro and in vivos. J Cell Mol Med. 2020 Dec;24(24):14217-14230. doi: 10.1111/jcmm.16038. Epub 2020 Oct 30. Erratum in: J Cell Mol Med. 2022 Jan;26(1):242-243. doi: 10.1111/jcmm.17117. PMID: 33128353; PMCID: PMC7754049.