KC-2741

293T-BA.1-SPIKE Cell Line

26781

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Background of 293T-BA.1-SPIKE Cell Line

BA.1, also known as S surface glycoprotein, is an immune escape variant. BA.1 is crucial for cell entry, pathogenicity, and is the key target of the adaptive immune response. BA.1 mediates attachment of the virus particle and entry into the host cell, and binds to the cellular protein angiotensin-converting enzyme 2 Subsequently, the BA.1 is primed by host cell proteases and drives the fusion of the viral and cellular membranes, which can take place at the plasma membrane or within endo-/lysosomes. BA.1 is an important target for vaccine development, antibody therapies and diagnostic antigen-based tests.

Specifications

Catalog Number	KC-2741
Cell Line Name	293T-BA.1-SPIKE Cell Line
Host Cell Line	293T
Description	Stable 293T cell line expressing exogenous BA.1 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 1:4-1:5 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
In Vivo Validation	NA

Cell Line Generation

293T-BA.1-SPIKE Cell Line was generated using a lentiviral vector expressing the BA.1 SPIKE sequence.

Characterization

Figure: Characterization of BA.1 overexpression in 293T stable clones using FACS.

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 1:4-1:5 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

BA.2 and BA.5 omicron differ immunologically from both BA.1 omicron and pre-omicron variants. Published: 13 December 2022. Nature Communications volume 13, Article number: 7701 (2022) Cite this article
Omicron spike protein: a clue for viral entry and immune evasion. Signal Transduction and Targeted Therapy volume 7, Article number: 339 (2022)
Spike (B.1.1.529 BA.1, Omicron Variant) (SARS-CoV-2) Pseudotyped Lentivirus (Luc Reporter)