KC-3768

Calu6-TROP2-Cell-Line

34742

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Background of Calu6-TROP2-Cell-Line

Trop2, also named as epithelial glycoprotein 1 antigen (EGP-1), is a member of a family including at least two type I membrane protein, Trop2 acts as a cell surface receptor and transduce intracellular calcium signal, and is also a carcinoma associated antigen, which was the target of several therapeutic antibodies, including GA773, and sacituzumab govitecan.

Specifications

Catalog Number	KC-3768
Cell Line Name	Calu6-TROP2-Cell-Line
Host Cell Line	Calu6
Description	Stable Calu6 clone expressing exogenous TROP2 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% RPMI 1640 + 20% FBS + 10% DMSO
Propagation Medium	RPMI 1640 + 10% FBS + 1μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Epithelial
Subculture	Split saturated culture 1:6-1:8 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 26 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

Calu6-TROP2-cell-line was generated using a lentiviral vector expressing the TROP2 sequence.

Characterization

Figure 1: Characterization of TROP2 overexpression in the Calu6-TROP2 stable clone using FACS.

Figure 2: Characterization of TROP2 in the Calu6-TROP2 stable clone using PCR sequencing.

Cell Resuscitation

1. Prewarm culture medium (RPMI 1640 supplemented with 10% FBS and 1μg/mL puromycin)in a 37°C water bath. 2. Thaw the frozen vial in a 37°C water bath for 1-2 minutes. 3. Transfer the vial into biosafety cabinet, and wipe the surface with 70% ethanol. 4. Unscrew the top of the vial and transfer the cell suspension gently into a sterile centrifuge tube containing 9.0mL complete culture medium. 5. Spin at ~ 125 × g for 5-7 minutes at room temperature, and discard the supernatant without disturbing the pellet. 6. Resuspend cell pellet with the appropriate volume of complete medium and transfer the cell suspension into a T25 culture flask. 7. Incubate the flask at 37°C, 5% CO₂ incubator. 8. Split saturated culture 1:6-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

1. Prepare the freezing medium (70% RPMI 1640 + 20% FBS + 10% DMSO) fresh immediately before use. 2. Keep the freezing medium on ice and label cryovials. 3. Transfer cells to a sterile, conical centrifuge tube, and count the cells. 4. Centrifuge the cells at 250×g for 5 minutes at room temperature and carefully aspirate off the medium. 5. Resuspend the cells at a density of at least 3×10⁶ cells/mL in chilled freezing medium. 6. Aliquot 1 mL of the cell suspension into each cryovial. 7. Freeze cells in the CoolCell freezing container overnight in a -80°C freezer. 8. Transfer vials to liquid nitrogen for long-term storage.

References

1.Linnenbach AJ, Wojcierowski J, Wu SA, et al. (1989). Sequence investigation of the major gastrointestinal tumor-associated antigen gene family, GA733. Proc. Natl. Acad. Sci. U.S.A. 86 (1): 27ÿ31 2. Fornaro M, Dell'Arciprete R, Stella M, et al. (1995). Cloning of the gene encoding Trop-2, a cell-surface glycoprotein expressed by human carcinomas. Int. J. Cancer. 62 (5): 610ÿ8. 3.El Sewedy T, Fornaro M, Alberti S (1998). Cloning of the murine TROP2 gene: conservation of a PIP2-binding sequence in the cytoplasmic domain of TROP-2. Int. J. Cancer. 75 (2): 324ÿ30.