Cx3cl1

Manufactured by R&D Systems

Sourced in United States, Austria

CX3CL1 is a lab equipment product offered by R&D Systems. It is a chemokine that functions as a cell adhesion molecule, mediating both cell-cell and cell-matrix interactions. The core function of CX3CL1 is to facilitate the migration and recruitment of leukocytes.

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Lab products found in correlation

Close spelling variants of this product

Human CX3CL1/Fractalkine Quantikine ELISA Kit (3 citations) Recombinant Rat CX3CL1 (2 citations) Anti-CX3CL1 (2 citations) Anti-CX3CL1 FITC (2 citations) Human CX3CL1/Fractalkine Quantikine ELISA (2 citations) Neutralizing antibody to CX3CL1 (2 citations) FITC anti-Human Fractalkine/CX3CL1 (2 citations) Fractalkine (CX3CL1) (2 citations) Recombinant human CX3CL1 (2 citations) Neutralizing antibody of CX3CL1 (2 citations)

24 protocols using cx3cl1

Fetal Resorption Analysis in Mice

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For IFN-γ treatment, homozygously mated BALB/c females were injected intraperitoneally with 5000 U IFN-γ (Peprotech, London, UK) or placebo (sodium phosphate containing 0.1% BSA) on GD6. For CX3CL1 treatment, homozygously mated BALB/c females were injected intraperitoneally with 1 μg of CX3CL1 (R&D Systems, Minneapolis, MN, USA) or placebo (PBS containing 0.1% BSA) on GD6. Mice were killed by cervical dislocation on GD8, and foetal resorption was assessed by observing the contents of uterus. Mice without gross implantation sites or tissue debris in the uterus were considered not pregnant and excluded from the experiment.
For histological analysis, uteri and ovaries were removed and fixed in 4% paraformaldehyde (PFA) overnight at 4 °C. After fixation, tissues were treated with ethanol and xylene and embedded in paraffin. Sections of 5 μm in thickness were prepared and stained using haematoxylin and eosin (H&E).

Li Z.Y., Chao H.H., Liu H.Y., Song Z.H., Li L.L., Zhang Y.J., Yang Y, & Peng J.P. (2014). IFN-γ induces aberrant CD49b+ NK cell recruitment through regulating CX3CL1: a novel mechanism by which IFN-γ provokes pregnancy failure. Cell Death & Disease, 5(11), e1512-.

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CX3CL1 Modulates Microglial Phagocytosis

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BV2 cells were cultured in Dulbecco's modified Eagle medium (DMEM) containing 10% fetal bovine serum (Gibco, USA) at 37 °C with 5% CO₂. For experiments, cells were seeded into six-well plates, and when confluence exceeded 60%, half of the wells were treated with CX3CL1 (30 ng/ml, R&D Systems), while the other half were treated with saline. The phagocytosis activity of BV2 cells that were either treated with CX3CL1 or untreated was assayed using microbeads with a diameter of 0.2 μm (Bio-Rad). Cells were incubated for 2 h with microbeads (1 μl), washed with PBS to eliminate free microbeads, and fixed onto glass slides using paraformaldehyde. Microglia were stained using an anti-Iba1 antibody (1:600; catalog no. ab5076, Abcam) for 24 h at room temperature, washed with PBS three times, and incubated with donkey anti-goat IgG (1:500; catalog no. 705-585-003, Jackson ImmunoResearch). Immunofluorescence images were obtained using a fluorescence microscope (Carl Zeiss) and analyzed using Image J.

Wang L., Ling H., He H., Hu N., Xiao L., Zhang Y., Xie L, & You Z. (2023). Dysfunctional synaptic pruning by microglia correlates with cognitive impairment in sleep-deprived mice: Involvement of CX3CR1 signaling. Neurobiology of Stress, 25, 100553.

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Pancreatic Cancer Cell Invasion and Migration Assay

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Cells were starved by serum free medium for 24 h prior to invasion and migration assays. Cell invasion and migration assays were both evaluated by the Boyden chamber method using the filter inserts (pore size, 8 μm), pre-coated (for invasion assay) or not pre-coated (for migration assay) with Matrigel in 24-well plates (BD Biosciences, Franklin Lakes, NJ, USA). Each pancreatic cancer cell line (7.5 × 10⁴ cells/insert for MIA PaCa-2, 2 × 10⁵ cells/insert for AsPC-1, or 4 × 10⁴ cells/insert for PANC-1) was seeded on the top chamber. The top chamber was filled with serum-free medium and the bottom chamber was filled with 10% FBS medium. The recombinant chemokines were then set in the bottom chamber at a final concentration of 25 ng/ml for CXCL8 (PeproTech, Rocky Hill, NJ, USA), and 0.5 μM for PGE2, 5 ng/mL for CX3CL1 (R&D Systems, Minneapolis, MN, USA) and C5a (ProSpec-Tany Technology Ltd, Ness Ziona, Israel). After incubation for 8 h in the case of PANC-1, or 48 h in the case of MIA PaCa-2 and AsPC-1cells, cells that passed through the filter and were attached to the lower surface of the filter were counted by staining with 0.01% crystal violet in 25% methanol. Cells attached to the lower surface of the filter membrane were then quantified by cell counting in five non-overlapping fields at × 100 magnification.

Saito K., Iioka H., Maruyama S., Sumardika I.W., Sakaguchi M, & Kondo E. (2019). PODXL1 promotes metastasis of the pancreatic ductal adenocarcinoma by activating the C5aR/C5a axis from the tumor microenvironment. Neoplasia (New York, N.Y.), 21(12), 1121-1132.

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Cytokine Profiling of Serum and Plasma

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Serum or plasma samples were immediately separated and kept at − 80 °C for subsequent cytokine analysis. Concentration of IL-6, IL-18, IL-17 (high sensitivity kit), IL-21, IL-22 (Affymetrix eBioscience, Vienna, Austria), IL-15, CCL20, CXCL13 and CX3CL1 (R&D Systems, Minneapolis, MN, USA) were measure by ELISA according to the manufacturer’s instructions.

Mangas-Losada A., García-García R., Leone P., Ballester M.P., Cabrera-Pastor A., Urios A., Gallego J.J., Martínez-Pretel J.J., Giménez-Garzó C., Revert F., Escudero-García D., Tosca J., Ríos M.P., Montón C., Durbán L., Aparicio L., Montoliu C, & Felipo V. (2019). Selective improvement by rifaximin of changes in the immunophenotype in patients who improve minimal hepatic encephalopathy. Journal of Translational Medicine, 17, 293.

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Ratiometric Imaging of Rac GTPase Activity

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For ratiometric imaging RAW/LR5 or fMLPR.2 cells were transiently transfected with Rac1 or Rac2 biosensor. Cells mounted on 12-mm round coverslips were serum-starved in RPMI for at least 1 h prior to stimulation with 50 ng/mL CX3CL1 (R&D Systems) or 100 nM fMLP (Sigma) in Buffer with Divalent (BWD = 125 mM NaCl, 5 mM KCl, 1 mM KH₂PO₄, 5 mM glucose, 10 mM NaHCO₃, 1 mM MgCl₂, 1 mM CaCl₂, and 20 mM Hepes) for indicated times at 37°C before fixation. Cells were fixed in 3.7% formaldehyde in BWD, permeabilized with 0.2% Triton X-100 in BWD for 10 min, stained for F-actin with Alexa Flour 568-phalloidin (1:400) (Invitrogen), and mounted in 50% glycerol in PBS. Microscope imaging at 60X magnification and image processing was performed as previously described in detail (24 (link), 37 (link), 38 (link)). Whole cell level of average GTPase activity was determined by thresholding the whole cell area in the FRET/donor ratiometric image using Metamorph Software. To measure GTPase activity at cell periphery we performed unbiased edge erosion measurements of ratio intensities as described below.

Miskolci V., Wu B., Moshfegh Y., Cox D, & Hodgson L. (2016). Optical tools to study the isoform-specific roles of small GTPases in immune cells. Journal of immunology (Baltimore, Md. : 1950), 196(8), 3479-3493.

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Cytokine and Chemokine Detection in Mice

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The cytokines and chemokines of the mice were detected using corresponding commercial ELISA kits. TNF-α (Cat#: MTA00B), IL-1β (Cat#: MLB00C), IL-6 (Cat#: M6000B), IL-18 (Cat#: DY122-05), MCP-1 (Cat#: SMJE00B), CCL1 (Cat#: DY845), IL-17 (Cat#: SM1700), IL-2 (Cat#: SM2000), IL-10 (Cat#: SM1000B), IL-5 (Cat#: SM5000), MIP-2 (Cat#: SMM200), MIP-3β (Cat#: DY440), MDC (Cat#: MCC220), CX3CL1 (Cat#: MCX310), CXCL10 (Cat#: DY466-05), and CXCL12 (Cat#: DY460) ELISA kits were purchased from R&D Systems (Shanghai, China) and used according to the product specifications. An MCP-3 (Cat#: BMS6006INST) ELISA kit was acquired from Invitrogen. A GCP-2 (Cat#: ab100719) kit was obtained from Abcam. An HMGB1 ELISA kit (Cat#: 326054329) was purchased from Shino-Test Corporation (Kanagawa, Japan). A procalcitonin ELISA kit (Cat#: EF014005) was obtained from Life Sciences. All the corresponding serum was carefully stored at −80 °C in a refrigerator until it was used.

Xu M., Ge C., Qin Y., Lou D., Li Q., Feng J., Wu Y., Hu L., Wang B, & Tan J. (2020). Functional loss of inactive rhomboid-like protein 2 mitigates obesity by suppressing pro-inflammatory macrophage activation-triggered adipose inflammation. Molecular Metabolism, 34, 112-123.

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Measuring Blood Ammonia and Cytokines

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Blood ammonia levels were measured immediately after blood collection using the Ammonia Test Kit II for the PocketChemBA system (Arkay, Inc., Kyoto, Japan). Plasma concentrations of IL-6, IL-18, IL-22 (Affymetrix eBioscience, Vienna, Austria), IL-15, CCL20, CXCL13 and CX3CL1 (R&D Systems, Minneapolis, MN, USA) were measured by ELISA according to the manufacturer’s instructions.

Casanova-Ferrer F., Gallego J.J., Fiorillo A., Urios A., Ríos M.P., León J.L., Ballester M.P., Escudero-García D., Kosenko E., Belloch V, & Montoliu C. (2024). Improved cognition after rifaximin treatment is associated with changes in intra- and inter-brain network functional connectivity. Journal of Translational Medicine, 22, 49.

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Comprehensive Murine and Human Chemokine Profiling

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Recombinant murine CCL2, 3, 4, 5, 6, 7, 8, 9/10, 11, 12, 19, 20, 21, 22, 24, 27, 28, CXCL1, 2, 4, 5, 9, 10, 11, 12 SDF1β, 13, 15, and 16 were purchased from PeproTech (Rocky Hill, NJ), and CCL17, 25, CXCL3, 17, CX3CL1, XCL1 and Chemerin were obtained from R&D Systems (Minneapolis, MN). In some cases, murine chemokines were not available, and human chemokines were used due to their known cross-species activity. Human CCL16, 18, CXCL6 and 8 were purchased from PeproTech, XCL2 was from Life Technologies (Carlsbad, CA) and CCL1, 13, 14, 15, 23, 26, CXCL7 and 14 were provided by ChemoCentryx (Mountain View, CA).

Yagawa Y., Robertson-Tessi M., Zhou S.L., Anderson A.R., Mulé J.J, & Mailloux A.W. (2017). Systematic Screening of Chemokines to Identify Candidates to Model and Create Ectopic Lymph Node Structures for Cancer Immunotherapy. Scientific Reports, 7, 15996.

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CX3CL1-CX3CR1 Axis Modulation

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PC-3 and VCaP cells were treated with CX3CL1 (100 nM; R&D Systems, Inc., Minneapolis, MN, USA) for 30 min at 37°C. For the lentiviral infections, PC-3 and VCaP cells (2×10⁵) were seeded onto 6-well plates. This was followed the next day by infection with either control or CX3CR1-overexpressing lentiviruses in the presence of polybrene (final concentration, 6 µg/µl). A total of 2–3 days following infection, cells were subcultured and selected with 5 µg/ml puromycin. For the siRNA transfections, PC-3 and VCaP cells (2×10⁵) were seeded onto 6-well plates, and 100 nM of either control siRNA or CX3CR1 siRNA was transfected using Lipofectamine^® 2000 reagent (Invitrogen; Thermo Fisher Scientific, Inc.). At 48 h post-transfection, the cells were recultured for 0–5 days for the different detection procedures. All the cells were divided into six groups, namely the blank control (CON), CX3CL1 group, overexpression negative control [NC (OE)], overexpression (OE), knockdown NC [NC (KD)] and knockdown (KD) groups, for cell functional analysis.

Liu P., Liang Y., Jiang L., Wang H., Wang S, & Dong J. (2018). CX3CL1/fractalkine enhances prostate cancer spinal metastasis by activating the Src/FAK pathway. International Journal of Oncology, 53(4), 1544-1556.

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Transwell Invasion Assay for Co-culture

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Transwell chambers with 8 μm pores (cat. no. 3402; Corning, Inc.) were used for assessing cell invasion. The upper chambers were pre-coated with Matrigel (cat. no. E6909; Sigma-Aldrich; Merck KGaA). For co-culture of the HCC cells and BMECs, the 24-well Transwell co-culture system was used. HCC cells at a density of 5×10⁴ cells and were plated in the upper chamber, while BMECs at a density of 2×10⁵ cells were seeded in the lower chamber. Both chambers were supplemented with serum-free medium. Subsequently, 100 nM CX3CL1 (R&D Systems, Inc.) or 50 ng/ml neutralizing antibody of CX3CL1 (cat. no. ab89229; Abcam) were added to the lower chamber. The cells were incubated at 37°C for 48 h, and the upper surfaces were gently swabbed to remove cells which had not migrated. The cells which had migrated to the lower membrane were fixed with 1% glutaraldehyde and stained with 0.1% crystal violet (cat. no. C0775; Sigma-Aldrich) for 15 min at room temperature. The number of cells which had invaded were counted in 5 randomly selected fields per well. Invasion experiments were performed 3 times independently.

Sun C., Hu A., Wang S., Tian B., Jiang L., Liang Y., Wang H, & Dong J. (2020). ADAM17-regulated CX3CL1 expression produced by bone marrow endothelial cells promotes spinal metastasis from hepatocellular carcinoma. International Journal of Oncology, 57(1), 249-263.

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