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Human CX3CR1-FLAG Stable Cell Line-HEK293T Gqi5

Cat.No.: CSC-RG0217
Cell Line Description: HEK293T Gqi5-HuCX3CR1-FLAG cell line is a hypotriploid human cell line, which has been transfected with a Human CX3C-chemokine receptor 1 (CX3CR1) tagged in the N-terminus with FLAG to allow stably express of the human CX3CR1 tagged in the N-terminus with FLAG protein. It is an example of a cell line transfected using our proprietary CBTGS gene screening and amplification system.
Background: CX3CR1, (CX3C-chemokine receptor 1, fractalkine receptor or GPR13) is a receptor for the CX3C chemokine fractalkine. CX3CR1 is expressed in cytotoxic effector lymphocytes, including natural killer cells, cytotoxic T lymphocytes and macrophages. Soluble fractalkine causes migration of these cells, whereas the membrane-bound form captures and enhances the subsequent migration in response to secondary stimulation with other chemokines. Furthermore, stimulation through membrane-bound fractalkine activates natural killer cells, leading to increased cytotoxicity and interferon-gamma production. Fractalkine is involved in the pathogenesis of various clinical disease states or processes, such as atherosclerosis, glomerulonephritis, cardiac allograft rejection and rheumatoid arthritis. In addition, polymorphisms in CX3CR1, which reduce its binding activity to fractalkine, have been reported to increase the risk of HIV disease and to reduce the risk of coronary artery disease.
Growth Properties: Adherent
Morphology: Epithelial
Host Cell: HEK293T Gqi5
Cell Line Validation: 1. Gene expression: RT-PCR experiments determined specific expression of human CX3CR1.2. Protein expression: CX3CR1 expression in this cell line has been validated by WB.3. Functional validation: Functional assays.
Sub-type: Chemokine
Propagation: Complete growth medium: DMEM, 10% FBS, 1 μg/mL puromycin, 250 μg/mL hygromycinAtmosphere: air, 95%; carbon dioxide (CO2), 5% Temperature: 37.0°C
Starting Cells From Frozen Cell Stock: 1. Remove the packaging cell lines from liquid nitrogen and carry out a quick thaw. Float the cells in the 37°C water bath for 2 minutes until nearly (80%) thawed. Once cells are thawed, it is important to dilute the cells 1:10 in growth media immediately to reduce the potentially toxic effects of the DMSO preservative on the cells.2. Clean the outside of the vial with 70% ethanol before opening.3. Remove the cells from the vial and add slowly into a 15ml conical tube containing 10 ml pre-warmed media. 4. Centrifuge for 3 minutes 1000 xg to pellet cells and remove the supernatant.5. Add 14 ml of media and transfer cells to a T25 flask or a 100 mm culture dish.6. Place the cells in the 37°C incubator with 5% CO2.7. Allow incubation for 3-4 days to reach confluence. The cells will re-attach to the surface over a period of several days in culture at 37°C.
Subculturing: 1. Remove and discard culture medium.2. Briefly rinse the cell layer with 0.25% (w/v) Trypsin-0.53 mM EDTA solution to remove all traces of serum that contains trypsin inhibitor.3. Add 2.0 to 3.0 ml of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 5 to 15 minutes).Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Cells that are difficult to detach may be placed at 37°C to facilitate dispersal.4. Add 6.0 to 8.0 ml of complete growth medium and aspirate cells by gently pipetting.5. Add appropriate aliquots of the cell suspension to new culture vessels.Incubate cultures at 37°C.Subcultivation Ratio: A subcultivation ratio of 1:4 to 1:10 is recommended Medium Renewal: 2 to 3 times per week
Mycoplasma: Mycoplasma Status: Negative (MycoAlert Kit)
Freeze Medium: Complete growth medium 90%; DMSO, 10%
Storage: Liquid nitrogen
Preservation: 1. Detach cells from culture dish according to the Sub-Culture Procedure.2. Resuspend cells at a density of 5 x 10^6 cells/mL in freeze medium.Note: A T-75 culture flask typically yields enough cells for preparing two frozen vials.3. Aliquot 1 mL cells into cryogenic vials.4. Place vials in a freezing container and store at –80 °C overnight.5. Transfer vials to liquid nitrogen for long term storage. If properly stored, cells should remain stable for years.
Safety Considerations: The following safety precautions should be observed.1. Use pipette aids to prevent ingestion and keep aerosols down to a minimum.2. No eating, drinking or smoking while handling the stable line.3. Wash hands after handling the stable line and before leaving the lab.4. Decontaminate work surface with disinfectant or 70% ethanol before and after working with stable cells.5. All waste should be considered hazardous.6. Dispose of all liquid waste after each experiment and treat with bleach.
Ship: Dry ice
Gene Name: CX3CR1 chemokine (C-X3-C motif) receptor 1 [ Homo sapiens ]
Official Symbol: CX3CR1
Synonyms: CX3CR1; chemokine (C-X3-C motif) receptor 1; chemokine (C X3 C) receptor 1 , CMKBRL1, GPR13; CX3C chemokine receptor 1; CCRL1; CMKDR1; V28; CMK-BRL1; CMK-BRL-1; C-X3-C CKR-1; fractalkine receptor; G protein-coupled receptor 13; G-protein coupled receptor 13; chemokine (C-X3-C) receptor 1; beta chemokine receptor-like 1; chemokine (C-C) receptor-like 1; GPR13; GPRV28; CMKBRL1;
Gene ID: 1524
mRNA Refseq: NM_001171171
Protein Refseq: NP_001164642
MIM: 601470
UniProt ID: P49238
Chromosome Location: 3p21.3
Pathway: Chemokine receptors bind chemokines, organism-specific biosystem; Chemokine signaling pathway, organism-specific biosystem; Chemokine signaling pathway, conserved biosystem; Class A/1 (Rhodopsin-like receptors), organism-specific biosystem; Cytokine-cytokine receptor interaction, organism-specific biosystem; Cytokine-cytokine receptor interaction, conserved biosystem; GPCR ligand binding, organism-specific biosystem;
Function: C-X3-C chemokine receptor activity; G-protein coupled receptor activity; chemokine receptor activity; protein binding; receptor activity; signal transducer activity;

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C-fusion    N-fusion   Non-tagged
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<1.0 eu/μg    <0.1 eu/μg   <0.01 eu/μg   Not required
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>80% by SDS-PAGE    >90% by SDS-PAGE   >95% by SDS-PAGE   Others

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