KC-5484

HT1080-NFκB-Luc2-LTBR-KO Cell Line

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Background of HT1080-NFκB-Luc2-LTBR-KO Cell Line

This gene encodes a member of the tumor necrosis factor receptor superfamily. The major ligands of this receptor include lymphotoxin alpha/beta and tumor necrosis factor ligand superfamily member 14. The encoded protein plays a role in signalling during the development of lymphoid and other organs, lipid metabolism, immune response, and programmed cell death. Activity of this receptor has also been linked to carcinogenesis. Alternatively spliced transcript variants encoding multiple isoforms have been observed.

Specifications

Catalog Number	KC-5484
Cell Line Name	HT1080-NFκB-Luc2-LTBR-KO Cell Line
Clone Number	3B2
Host Cell Line	HT1080
Description	Stable HT1080-NFκB-Luc2 clone with human LTBR gene knockout, No.3B2
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+150μg/mL Hygromycin B
Selection Marker	Hygromycin B
Morphology	epithelial
Subculture	Split saturated culture 1:6-1:10 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

HT1080-NFκB-Luc2-LTBR-KO cell line was generated using the CRISPR method.

Characterization

Figure 1: Characterization of HT1080-NFκB-Luc2-LTBR-KO cell line stable clone using PCR sequencing.

Figure 2: Characterization of HT1080-NFκB-Luc2-LTBR-KO cell line stable clone using RT-PCR sequencing.

Figure 3: Characterization of HT1080-NFκB-Luc2-LTBR-KO cell line stable clone using Western blot.

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 1:6-1:10 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

https://www.ncbi.nlm.nih.gov/gene/4055