KC-2494

293T-CCR2b-High-Cell-Line

24174

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Background of 293T-CCR2b-High-Cell-Line

The chemokine (C-C motif) receptor 2B (CCR2B) is one of the two isoforms of the receptor for monocyte chemoattractant protein-1 (CCL2), are present that belong to the rhodopsin-like G protein-coupled receptor family, and couple to Gi and Gq family members. CCR2 has been found to be expressed by multiple cell types including monocytes, dendritic cells (DCs), endothelial cells, certain T cell subsets, and cancer cells. CCR2 mediates agonistdependent calcium mobilization and inhibition of adenylyl cyclase, and it can also be a coreceptor with CD4 for HIV1 infection. CCL2-CCR2 signaling axis is implicated in many inflammatory and neurodegenerative diseases such atherosclerosis, multiple sclerosis, asthma, neuropathic pain, diabetic nephropathy, and cancer. PF-4136309 is an inhibitor of CCR3.

Specifications

Catalog Number	KC-2494
Cell Line Name	293T-CCR2b-High-Cell-Line
Host Cell Line	Human HEK293T cell line
Description	Stable HEK293T cell line expressing exogenous human CCR2b gene
Quantity	One vial 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 1:3-1:6 every 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 human CCR2b cell line was generated using a lentiviral vector expressing the human CCR2b sequence.

Characterization

Cell Resuscitation

Prewarm culture medium (DMEM + 10% FBS + 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 1:3-1:6 every 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

Minsaas L, Planagum½Ç J, Madziva M, Krakstad BF, Masi½Ç-Balagu¹È M, Katz AA, Aragay AM. Filamin a binds to CCR2B and regulates its internalization. PLoS One. 2010 Aug 17;5(8):e12212. doi: 10.1371/journal.pone.0012212. PMID: 20808917; PMCID: PMC2923182.
Markx D, Schuhholz J, Abadier M, Beier S, Lang M, Moepps B. Arginine 313 of the putative 8th helix mediates GÏ«q/14 coupling of human CC chemokine receptors CCR2a and CCR2b. Cell Signal. 2019 Jan;53:170-183. doi: 10.1016/j.cellsig.2018.10.007. Epub 2018 Oct 12. PMID: 30321592.
Hao Q, Vadgama JV, Wang P. CCL2/CCR2 signaling in cancer pathogenesis. Cell Commun Signal. 2020 May 29;18(1):82. doi: 10.1186/s12964-020-00589-8. PMID: 32471499; PMCID: PMC7257159.

Use License Agreement

Research Use Only.

Not for use in diagnostic procedures or therapeutic applications.

Redistribution of the cell line or its derivatives is prohibited without prior written permission from Kyinno Biotechnology.