KC-5106

HeLa-ERBB2-KO Cell Line

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Background of HeLa-ERBB2-KO Cell Line

The ERBB2 gene, also known as HER2, encodes a tyrosine kinase receptor belonging to the epidermal growth factor receptor (EGFR) family. This protein has no ligand binding domain of its own and therefore cannot bind growth factors. However, it does bind tightly to other ligand-bound EGF receptor family members to form a heterodimer, stabilizing ligand binding and enhancing kinase-mediated activation of downstream signalling pathways, such as those involving mitogen-activated protein kinase and phosphatidylinositol-3 kinase. Amplification and/or overexpression of this gene has been reported in numerous cancers, including breast and ovarian tumors. Alternative splicing results in several additional transcript variants, some encoding different isoforms and others that have not been fully characterized.

Specifications

Catalog Number	KC-5106
Cell Line Name	HeLa-ERBB2-KO Cell Line
Clone Number	3C1
Host Cell Line	HeLa
Description	Stable HeLa clone with human ERBB2 gene knockout, No.3C1
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	DMEM+20% FBS+10% DMSO
Propagation Medium	DMEM+10% FBS
Selection Marker	NA
Morphology	epithelial
Subculture	Split saturated culture 1:5-1:8 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 40 hours
Mycoplasma Status	Negative

Cell Line Generation

HeLa-ERBB2-KO cell line was generated using the CRISPR method.

Characterization

Figure 1: Characterization of HeLa-ERBB2-KO-3C1 cell line stable clone using PCR sequencing.

Figure 2: Characterization of HeLa-ERBB2-KO-3C1 cell line stable clone using RT-PCR sequencing.

Figure 3: Characterization of HeLa-ERBB2-KO-3C1 cell line stable clone using FACS.

Cell Resuscitation

Prewarm culture medium (DMEM + 10% FBS) 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 1:5-1:8 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

Pilati C, Soulabaille A, Gallois C, Blons H, Cayre A, Sroussi M, Le Corre D, Mouillet-Richard S, Mulot C, Le Malicot K, De Reynies A, Bachet JB, Borg C, Di Fiore F, Guimbaud R, Bennouna J, André T, Taieb J, Penault-Llorca F, Laurent-Puig P. ERBB2 Comprehensive Profiling and Prognostication in Stage III Colon Cancer: Findings From PETACC8 and IDEA-France Cohorts. Gastroenterology. 2025 Apr;168(4):714-724.e4. doi: 10.1053/j.gastro.2024.10.046. Epub 2024 Nov 28. PMID: 39612956.
https://www.ncbi.nlm.nih.gov/gene/2064