KC-5965

293T-ITGA4-ITGB7 Cell Line

63009

Home » 293T-ITGA4-ITGB7 Cell Line

Background of 293T-ITGA4-ITGB7 Cell Line

ITGA4 (Integrin Subunit Alpha 4), also known as CD49d, and ITGB7 (Integrin Subunit Beta 7) form the heterodimeric integrin α4β7. This receptor is primarily expressed on subsets of lymphocytes and mediates the gut-specific homing of T cells by binding to MAdCAM-1 on intestinal endothelium. Aberrant expression of α4β7 is linked to inflammatory bowel disease (IBD), where it drives pathological lymphocyte infiltration into the gut mucosa, and also plays a role in HIV pathogenesis by facilitating viral spread and intestinal homing. Drug development has yielded vedolizumab, an FDA-approved monoclonal antibody that blocks α4β7-MAdCAM-1 interaction for the treatment of ulcerative colitis and Crohn’s disease. Additionally, natalizumab targets the α4 subunit, blocking both α4β1 and α4β7, and is approved for multiple sclerosis and Crohn’s disease.

Specifications

Catalog Number	KC-5965
Cell Line Name	293T-ITGA4-ITGB7 Cell Line
NCBI/UniProt Accession Number	NM_000885, NM_000889
Clone Number	25#
Host Cell Line	293T
Description	Stable 293T cell line expressing exogenous human ITGA4 and ITGB7 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% basal medium+20% FBS+10% DMSO
Propagation Medium	DMEM + 10% FBS + 1μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Epithelial-like
Subculture	Split saturated culture 1:4-1:8 every 2-3 days
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-ITGA4-ITGB7 cell line was generated using a lentiviral vector expressing the ITGA4 and ITGB7 sequence.

Characterization

Figure 1: Characterization of ITGA4-ITGB7 overexpression in the 293T-ITGA4-ITGB7 stable clone using FACS.

Figure 2: Characterization of ITGB7 overexpression in the 293T-ITGA4-ITGB7 stable clone using FACS.

Figure 3: Characterization of ITGA4 and ITGB7 in the 293T-ITGA4-ITGB7 stable clone using PCR sequencing.

Cell Resuscitation

Pre-warm complete culture medium (basal medium and 10% FBS) in a 37°C water bath.
Rapidly thaw the cryovial in a 37°C water bath for 1-2 minutes with gentle agitation.
Transfer the vial to a biosafety cabinet, and disinfect the exterior with 70% ethanol.
Aseptically transfer the cell suspension dropwise into a sterile centrifuge tube containing 9.0 mL of pre-warmed complete medium.
Centrifuge at approximately 125 × g for 5–7 minutes at room temperature, carefully aspirate the supernatant without disturbing the cell pellet.
Gently resuspend the pellet in an appropriate volume of complete medium and transfer the suspension into a T25 flask.
Incubate the flask in a 37°C in a humidified 5% CO₂ incubator.
Assess cell viability and morphology after 24 hours. If cells appear healthy, replace the medium with fresh medium supplemented with the appropriate selective antibiotic.
Subculture the cells at a ratio of 1:4-1:8 every 2-3 days upon reaching 80%–90% confluency.

Cell Freezing

Prepare the freezing medium (70% basal medium, 20% FBS and 10% DMSO) freshly before use.
Pre-chill the freezing medium on ice and label the cryovials accordingly.
Transfer the cell suspension to a sterile conical tube and perform a cell count to determine total viability and density.
Centrifuge the cells at 250×g for 5 minutes at room temperature; carefully aspirate the supernatant.
Gently resuspend the cell pellet in chilled freezing medium, ensuring a minimum cell density of 3×10⁶ cells/mL.
Aliquot 1 mL of the cell suspension into each pre-labeled cryovial.
Place the cryovials into a CoolCell® container and store at -80°C overnight for controlled-rate cooling.
Transfer the cryovials to the liquid nitrogen for long-term storage the following day.

References

1. Cushing, Kelly, and Peter D R Higgins. “Management of Crohn Disease: A Review.” JAMA vol. 325,1 (2021): 69-80. doi:10.1001/jama.2020.18936.
2. Gros, Beatriz, and Gilaad G Kaplan. “Ulcerative Colitis in Adults: A Review.” JAMA vol. 330,10 (2023): 951-965. doi:10.1001/jama.2023.15389.
3. Slack, R J et al. “Emerging therapeutic opportunities for integrin inhibitors.” Nature reviews. Drug discovery vol. 21,1 (2022): 60-78. doi:10.1038/s41573-021-00284-4.