Il 6 elisa kit

Manufactured by R&D Systems

Sourced in United States, United Kingdom, Germany

The IL-6 ELISA kit is a laboratory test used to measure the concentration of interleukin-6 (IL-6) in biological samples. IL-6 is a cytokine that plays a role in immune responses and inflammatory processes. The kit utilizes the enzyme-linked immunosorbent assay (ELISA) technique to detect and quantify IL-6 levels in the sample.

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Close spelling variants of this product

ELISAs (144 citations) IL-6 kits (2 citations)

88 protocols using il 6 elisa kit

Extraction of Inflammatory Proteins

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To extract proteins from tissues of 0.5% λ-carrageenan-induced paw edema models, the tissues were frozen in liquid nitrogen, pulverized using a mortar, and lysed in PRO-PREP™. The lysed tissue was centrifuged at 13,000 rpm for 15 min at 4 °C to separate the supernatant. The separated supernatant was subjected to ELISA in accordance with the manufacturer’s protocol to confirm and measure iNOS, COX-2, IL-1β, and IL-6 levels using the following kits: iNOS ELISA kit (MBS261100, mybiosource, BC, CA), COX-2 ELISA kit (MBS269104, mybiosource, BC, CA), IL-1β ELISA kit (MLB00C, R&D Systems, USA), IL-6 ELISA kit (M6000B, R&D Systems, USA).

Kim J.H., Hong M., Han J.H., Ryu B.R., Lim Y.S., Lim J.D., Kim C.H., Lee S.U, & Kwon T.H. (2023). In Vitro and In Vivo Anti-Inflammatory Effects of Cannabidiol Isolated from Novel Hemp (Cannabis sativa L.) Cultivar Pink Pepper. Molecules, 28(18), 6439.

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IL-6 Measurement in Cell Culture

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Cells were incubated with 1 mL of RPMI medium with 10% FCS in a 24-well-plate (2 × 10⁴ cells/well) for a period of 36 h. After incubation, we collected supernatants from each well and measured IL-6 levels in the conditioned media by an IL-6 ELISA kit according to manufacturer’s instructions (Cat No. D6050; R & D Systems, Inc.). The IL-6 levels were adjusted by the concentrations of protein in the whole cell extract, which was measured using a BCA protein assay kit (Pierce, Rockford, IL, USA), as previously described [28 (link)].

Chiang K.C., Chang K.S., Hsu S.Y., Sung H.C., Feng T.H., Chao M, & Juang H.H. (2020). Human Heme Oxygenase-1 Induced by Interleukin-6 via JAK/STAT3 Pathways Is a Tumor Suppressor Gene in Hepatoma Cells. Antioxidants, 9(3), 251.

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IL-6 Secretion Measurement in IHOK Cells

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Each IHOK cell line was cultured with proper culture medium for 48 hr in a 100 mm dish. Their conditioned media were harvested for the analysis of IL‐6 secretion. Concentration of IL‐6 was measured according to the manufacturer's protocol using IL‐6 ELISA kit (R&D, Minneapolis, MN).

Son H., Park I., Kim J.Y., Kim D.K., Illeperuma R.P., Bae J.Y., Lee D.Y., Oh E., Jung D., Williams D.R, & Kim J. (2015). A Distinct Role for Interleukin‐6 as a Major Mediator of Cellular Adjustment to an Altered Culture Condition. Journal of Cellular Biochemistry, 116(11), 2552-2562.

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Cytokine Profiling of Conditioned Media

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Conditioned medium was collected after 24 h incubation from confluent cells and was applied to the Human Cytokine Antibody Array C1000 (Raybiotech) or IL6 ELISA kit (R&D) following the manufacturer’s instructions.

Wang J., Rouse C., Jasper J.S, & Pendergast A.M. (2016). ABL kinases promote breast cancer osteolytic metastasis by modulating tumor-bone interactions through TAZ and STAT5 signaling. Science signaling, 9(413), ra12.

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Quantification of IL6 and OSM in Cell Lines

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HBEC3K, NLF, and CAF cells were seeded at 2×10⁵ cells per 60-mm cell culture dish and incubated for 72 hours. For measurement of IL6 concentration, cell culture media were collected and subjected to an IL6 ELISA kit (R&D Systems) according to the manufacturer’s instructions. For measurement of OSM, cells were subjected to lysis in cell lysis buffer (Sigma) analyzed with an OSM ELISA kit (R&D Systems).

Shien K., Papadimitrakopoulou V.A., Ruder D., Behrens C., Shen L., Kalhor N., Song J., Lee J.J., Wang J., Tang X., Herbst R.S., Toyooka S., Girard L., Minna J.D., Kurie J.M., Wistuba I.I, & Izzo J.G. (2017). JAK1/STAT3 activation through a proinflammatory cytokine pathway leads to resistance to molecularly targeted therapy in non-small cell lung cancer. Molecular cancer therapeutics, 16(10), 2234-2245.

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Intercellular signaling and cell migration

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In vitro intercellular cell signaling assay was performed using macrophages (J774, ATCC) and fibroblasts (3T3, ATCC). Briefly, macrophages alone, fibroblasts alone or co-cultured macrophages and fibroblasts were cultured in 24 well plates and grown to 80% confluence followed by supplementing 10ng/mL of MCP-1 protein (R&D Systems) or control PBS. Forty-eight hours later, culture medium was collected and IL-6 levels were measured using an IL-6 ELISA kit (R&D Systems).
The recruitment effect of IL-6 on ECs (HUVEC, C-003-5C, Invitrogen), macrophages (J774, ATCC), SMCs (MOVAS, ATCC), and fibroblasts (3T3, ATCC) was studied in vitro by cell migration assay. Oris Pro Cell Migration Assay kit was used for cell migration assay. All cells were cultured with media supplemented with fetal bovine serum (FBS) and grown to 100% confluence followed by serum starvation for 12 hours in 96-well cell culture plates. The assay was started with 50 ng/mL of IL-6 protein or PBS supplemented in culture medium. All procedures followed the manufacturer’s instructions. Twenty-four hours after the assay started, cell migratory area was measured using Image Pro software.

Hosaka K., Rojas K., Fazal H.Z., Schneider M.B., Shores J., Federico V., McCord M., Lin L, & Hoh B. (2017). The Monocyte Chemotactic Protein-1 — Interleukin-6 — Osteopontin Pathway of Intra-Aneurysmal Tissue Healing. Stroke, 48(4), 1052-1060.

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Radiation-Induced IL-6 Secretion Assay

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Supernatant from SUM159PT cells either untreated or treated with increasing doses of radiation were assayed with an IL-6 ELISA kit (R&D Systems) as described in the manufacturer’s protocol. Supernatants were collected 24, 48 or 72 hours post-irradiation and centrifuged to remove cells and debris. In addition, cells were trypsinized and counted by trypan blue exclusion using a Countess automated cell counter (Invitrogen) to normalize secreted IL-6 to total cell number. IL-6 ELISA was performed according to the manufacturer’s instructions. Briefly, 100 μl of supernatant or supplied standard was added to each well containing 100 μl of assay diluent and incubated at room temperature for 2 hours. After four washes, 200 μl of conjugate was added to each well and incubated at room temperature for an additional 2 hours. Following four additional washes, 200 μl of substrate was added to each well and incubated in the dark for 20 minutes. Finally, 50 μl of stop solution was added to each well and the absorbance was read at 450 nm. The standards were graphed and used to determine the concentration of IL-6 in each of the supernatant samples.

Arnold K.M., Opdenaker L.M., Flynn N.J., Appeah D.K, & Sims-Mourtada J. (2020). Radiation induces an inflammatory response that results in STAT3-dependent changes in cellular plasticity and radioresistance of breast cancer stem-like cells. International journal of radiation biology, 96(4), 434-447.

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Cardiac Injury Assessment in Mice

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Blood samples were collected from the mice to detect cardiac troponin I (cTnI) and N-terminal probrain natriuretic peptide (NT-proBNP) for the evaluation of cardiac injury. Mouse NT-proBNP (#CSB-E05153m) kit and cTnI ELISA kit (#CSB-E08421m) were obtained from CUSABIO (Wuhan, China).
The superoxide dismutase (SOD) activity assay kit, catalase activity kit, glutathione peroxidase (Gpx) activity kit, and glutathione (GSH) content assay kit were all purchased from Naijing Jiancheng Bioengineering Institute (Nanjing, China). Malondialdehyde (MDA) assay kit was used to detect MDA in the supernatant via thiobarbituric acid method according to the instructions. 4-Hydroxynonenal (4-HNE)-protein adducts detection kit was provided by Abcam (#ab238538). 3-Nitrotyrosine (3-NT) competitive ELISA was provided by Abcam (#ab113848).
DNA-p65 nuclear factor kappa-B (NF-κB) binding assay was detected with a Mercury TransFactor kit (BD Biosciences, Clontech). To detect myocardial inflammatory response, fresh heart samples were homogenized to detect myocardial tumor necrosis factor (TNF)-α and interleukin (IL)-6 expression. TNF-α Mouse ELISA Kit and IL-6 ELISA Kit were provided by R&D Systems.

Zhang W.B., Zheng Y.F, & Wu Y.G. (2021). Inhibition of miR-128-3p Attenuated Doxorubicin-Triggered Acute Cardiac Injury in Mice by the Regulation of PPAR-γ. PPAR Research, 2021, 7595374.

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Biochemical Assays and Cytokine Quantification

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RES raw powders were purchased from Sigma Chemical Co. (St. Louis, MO, USA), while ISO injection was obtained from a local pharmacy. L-RES was purchased from Lipolife, UK. L-RES has a particle size less than 200 nm with a neutral zeta potential as stated by the manufacturer. Urea, creatinine and uric acid biochemical assays were supplied from Randox, Crumlin, UK. IL-6 ELISA kit was obtained from R&D Systems (Minneapolis, MN, USA). MAPK, cystatin c and β-actin antibodies were purchased from Novus Biologicals (Centennial, CO, USA). The PCR primers were obtained from Sigma (St. Louis, MO, USA).

Alhusaini A.M., Alshehri S.M., Sarawi W.S., Alghibiwi H.K., Alturaif S.A., Al khbiah R.A., Alali S.M., Alsaif S.M., Alsultan E.N, & Hasan I.H. (2023). Implication of MAPK, Lipocalin-2, and Fas in the protective action of liposomal resveratrol against isoproterenol-induced kidney injury. Saudi Pharmaceutical Journal : SPJ, 32(1), 101907.

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Alisol F and 25-anhydroalisol F Isolation and Effects

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Alisol F and 25-anhydroalisol F were isolated from A. orientale by one of the authors (Q. Ma) using our previously-established method and provided with a purity of 98.0% determined by high-pressure liquid chromatography. Dulbecco’s Modified Eagle Medium (DMEM), fetal bovine serum (FBS), 0.25% trypsin, and penicillin-streptomycin-amphotericin (PSA) were purchased from Gibco BRL Co. Ltd. (Gaithersbug, MD, USA). Lipopolysaccharide (LPS) from Escherichia coli 055:B5, 3-[4,5-Dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT), dexamethasone (DXM), d-galactosamine (d-gal) and Trizol reagent were obtained from Sigma Chemical Co. (St. Louis, MO, USA). The Griess reagent, the protein extraction kit and BCA protein assay kit were obtained from Beyotime Institute of Biotechnology (Beijing, China). Go Tag^® qPCR Master Mix and GoScript^TM Reverse Transcription System were purchased from Promega (Madison, WI, USA). Rabbit polyclonal antibodies against inducible nitric oxide synthase (iNOS), COX-2, p-p38 (Thr180/Tyr182), p38, p-ERK1/2 (Thr202/Try204), ERK1/2, p-SAPK/JNK (Thr183/Tyr185), SAPK/JNK, p-p65 (Ser536), p65, GAPDH, p-IκB-α (Ser32), IκB-α, p-STAT3 (Tyr705), STAT3, goat-IgG HRP, and lamin B1 were purchased from Cell Signaling Technology (Danvers, MA, USA). Mouse TNF-α, IL-1β, and IL-6 ELISA kit were from R and D systems (Abingdon, UK).

Bi X., Wang P., Ma Q., Han L., Wang X., Mu Y., Guan P., Qu X., Wang Z, & Huang X. (2017). Anti-Inflammatory Activities and Liver Protection of Alisol F and 25-Anhydroalisol F through the Inhibition of MAPK, STAT3, and NF-κB Activation In Vitro and In Vivo. Molecules : A Journal of Synthetic Chemistry and Natural Product Chemistry, 22(6), 951.

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