KC-2389

293T-cyno-CXCR5-Cell-Line

23960

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Background of 293T-cyno-CXCR5-Cell-Line

CXCR5, also known as CD185, encodes a multi-channel membrane protein belonging to the CXC chemokine receptor family. It is selectively expressed in B cells and participates in lymphocyte homing and the development of normal lymphoid tissue. CXCL13 and its receptor CXCR5 are associated with the pathogenesis of various autoimmune diseases, such as rheumatoid arthritis, multiple sclerosis, systemic lupus erythematosus, primary Sjogren's syndrome, myasthenia gravis, and inflammatory bowel disease.

Specifications

Catalog Number	KC-2389
Cell Line Name	293T-cyno-CXCR5-Cell-Line
Host Cell Line	293T
Description	Stable 293T clone expressing exogenous cyno CXCR5 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 1:4-1:8 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 30 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

293T-cyno-CXCR5-cell-line was generated using a lentiviral vector expressing the cyno CXCR5 sequence.

Characterization

Figure 1: Characterization of cyno CXCR5 overexpression in the 293T-cyno-CXCR5 stable clone 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 1:4-1:8 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

Makhlouf MM, Radwan ER, Khorshed OM, Fathi LM, Elmasry MM. CXC Chemokine Receptor Type 5 Gene Polymorphisms in a Cohort of Egyptian Patients with Diffuse Large B-Cell Lymphoma. Pathobiology. 2021;88(3):211-217. doi: 10.1159/000510456. Epub 2020 Dec 30. PMID: 33378752.
Pan Z, Zhu T, Liu Y, Zhang N. Role of the CXCL13/CXCR5 Axis in Autoimmune Diseases. Front Immunol. 2022 Mar 4;13:850998. doi: 10.3389/fimmu.2022.850998. Erratum in: Front Immunol. 2022 Oct 20;13:1061939. PMID: 35309354; PMCID: PMC8931035.