KC-5400

Ba/F3-ERBB3 Cell Line

58290

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Background of Ba/F3-ERBB3 Cell Line

ERBB3, also known as HER3, is a transmembrane protein belonging to epidermal growth factor receptor (ERBB) family of receptor tyrosine kinase. The kinase domain impaired ERBB3 can from active heterodimer with other member of ERBB family, this heterodimer, mostly with ERBB2 after binding with its ligand NRG1 or NRG2, play an important role in cancer growth, invasion, metastasis and chemotherapeutic resistant.

Specifications

Catalog Number	KC-5400
Cell Line Name	Ba/F3-ERBB3 Cell Line
Clone Number	12#
Host Cell Line	Ba/F3
Description	Stable Ba/F3 clone expressing exogenous ERBB3 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% RPMI1640+20% FBS+10% DMSO
Propagation Medium	RPMI1640+10%FBS+8ng/mL mIL3+1000μg/mL G418
Selection Marker	G418
Morphology	Mostly single, round (some polymorph) cells in suspension
Subculture	Split saturated culture 1:10 every 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 20 hours
Mycoplasma Status	Negative

Cell Line Generation

Ba/F3 ERBB3 cell line was generated using retrovirus vector expressing ERBB3 gene.

Characterization

Figure 1: Characterization of ERBB3 overexpression in the Ba/F3 ERBB3 stable clone using PCR sequence.

Figure 2: Characterization of ERBB3 overexpression in the Ba/F3 ERBB3 stable clone using FACS.

Cell Resuscitation

1. Prewarm culture medium (RPMI1640+10%FBS+8ng/mL mIL3+1000μg/mL G418)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 1:10 every 3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

1. Prepare the freezing medium (70% RPMI1640 + 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. Jaiswal, B. S. et al. Oncogenic ERBB3 Mutations in Human Cancers. Cancer Cell 23, 603ÿ617 (2013).
2.Baselga, J. & Swain, S. M. Novel anticancer targets: Revisiting ERBB2 and discovering ERBB3. Nat. Rev. Cancer 9, 463ÿ475 (2009).