Mab208

Manufactured by R&D Systems

Sourced in United States

MAB208 is a monoclonal antibody that recognizes human CD163. CD163 is a scavenger receptor expressed on the surface of monocytes and macrophages. This antibody is useful for the identification and characterization of CD163-positive cells.

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Lab products found in correlation

21 protocols using mab208

Neutralization Assay for CXCL1, IL-8, and IL-18

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The CXCL1 antibody (9µg/mL; cat. no. MAB275; R&D SYSTEMS, Minneapolis, MN, USA), IL-8/CXCL8 (0.4µg/mL; cat. no. MAB208; R&D SYSTEMS, Minneapolis, MN, USA), and IL-18 (1.2µg/mL; cat. no. AF2548; R&D SYSTEMS, Minneapolis, MN, USA) were used for the neutralization assay. The normal mouse IgG1 antibody (1 µg/mL; cat. no. sc-3877; Santa Cruz Biotechnology, Inc., Dallas, TX, USA) was used as the control antibody. The experiment was conducted in the same manner as the cell culture environment, and the subsequent experiments or results were performed/obtained after 24–48 h.

Kahm Y.J., Kim I.G, & Kim R.K. (2023). RanBP1: A Potential Therapeutic Target for Cancer Stem Cells in Lung Cancer and Glioma. International Journal of Molecular Sciences, 24(7), 6855.

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Quantification of sCD14 and CXCL8 in cell supernatants

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Soluble human CD14 (sCD14) was measured in the supernatants with a purchased DuoSet ELISA CD14 kit (R&D Systems, Europe, Abingdon, UK). The analysis was performed according to manufacturers’ protocol and the detection range was 62.5–4,000 pg/ml. For duplicate samples an intra-assay coefficient of variation (CV)<10% was accepted.
Interleukin-8 (CXCL8) was measured in the supernatants with in-house ELISA methods (14 (link)). Commercially available antibody pair MAB 208 and BAF 208 for detection of CXCL-8 (R&D systems, Europe, Abingdon, UK) was used as previously described. The detection range was 40–3,200 pg/ml. For duplicate samples, an intra-assay CV <10% was accepted.

von Schéele I., Larsson K, & Palmberg L. (2014). Interactions between alveolar epithelial cells and neutrophils under pro-inflammatory conditions. European Clinical Respiratory Journal, 1, 10.3402/ecrj.v1.24545.

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Erlotinib and Anti-IL-8 Antibody Effects

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Tumor cells were pre-treated for 72 h with 100 nM erlotinib as indicated above, or left untreated. A third group consisted of tumor cells treated with erlotinib for 72 h followed by treatment with a commercially available neutralizing anti-IL-8 antibody (MAB208, R&D Systems, Minneapolis, MN, USA; 10 μg/ml) for 96 h. Cells were labeled and used as targets with NK effector cells or recombinant TRAIL.

Dominguez C., Tsang K.Y, & Palena C. (2016). Short-term EGFR blockade enhances immune-mediated cytotoxicity of EGFR mutant lung cancer cells: rationale for combination therapies. Cell Death & Disease, 7(9), e2380-.

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Cytokine Detection in Cell Supernatant

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Detection of interleukin-8/CXCL8 in the supernatant was performed using an in-house enzyme-linked immunosorbent assay (ELISA) method.15 (link) Commercially available antibody pair MAB 208 and BAF 208 (R&D Systems, Inc.) was used to detect IL-8. The detection range for IL-8 was 12.5–6,400 pg/mL. The measurements of TNF-α in the supernatant were performed by using high-sensitive Quantikine ELISA kit (R&D Systems, Inc.). The analyses of TNF-α were performed according to the protocol from the manufacturer. For all the duplicated samples, an intra-assay variation <10% (for TNF-α, <20%) was accepted.

Ji J., von Schéele I., Billing B., Dahlén B., Lantz A.S., Larsson K, & Palmberg L. (2016). Effects of budesonide on toll-like receptor expression in alveolar macrophages from smokers with and without COPD. International Journal of Chronic Obstructive Pulmonary Disease, 11, 1035-1043.

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Modulation of Wnt and Cytokine Signaling

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Cell lines and patient derived samples were treated with the following prior to downstream assays: 50 ng/ml DKK1 (GF170, Milipore), 50 µg/ml Vantictumab (Oncomed), 100 nm Wnt3A (5036-WN, R&D systems), 100 µM LGK974 (S7143, Selleckchem), 10 ng/ml rIL15 (247-ILB, R&D systems), 10 ng/ml rIL1β (201-LB, R&D systems), 5 µg/ml IL1β neutralising antibody (MAB201, R&D systems), 5 µg/ml IL15 neutralising antibody (MAB-274, R&D systems), 5 µg/ml IL6 neutralising antibody (MAB2061, R&D systems), 5 µg/ml IL8 neutralising antibody (MAB208, R&D systems), 10 µg/ml Anakinra (Amgen, Cambridge, UK), 5 mM Sulfasalazine (Sigma), 10uM KG-501 (Sigma). Treatment times for individual assays are detailed below.

Eyre R., Alférez D.G., Santiago-Gómez A., Spence K., McConnell J.C., Hart C., Simões B.M., Lefley D., Tulotta C., Storer J., Gurney A., Clarke N., Brown M., Howell S.J., Sims A.H., Farnie G., Ottewell P.D, & Clarke R.B. (2019). Microenvironmental IL1β promotes breast cancer metastatic colonisation in the bone via activation of Wnt signalling. Nature Communications, 10, 5016.

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Reagents for Cytokine Quantification

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Monoclonal human IL8/CXCL8 antibody (anti-hIL8, MAB208), recombinant human IL8/CXCL8 protein (hIL8, 208-IL), mouse TNFα antibody (MAB410), recombinant mouse TNF-α protein (410-MT), mouse IL6 antibody (MAB406), recombinant mouse IL6 protein (406 ML), and CXCL8/IL8 Alexa Fluor 488-conjugated antibody (IC208G) were purchased from R&D Systems (Minneapolis, MN, USA). A Sylgard 184 silicone elastomer kit was purchased from Dow Corning. Phosphate-buffered saline at 1× concentration (PBS, pH = 7.4) was purchased from Sigma-Aldrich (MO, USA). All other chemicals were of analytical grade.

Song N., Xie P., Shen W., Oh H., Zhang Y., Vitale F., Javanmard M, & Allen M.G. (2021). A microwell-based impedance sensor on an insertable microneedle for real-time in vivo cytokine detection. Microsystems & Nanoengineering, 7, 96.

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Quantifying CXCL8 and CCL3 in Monocytes

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Quantification of CXCL8 in monocyte supernatants was done by ELISA. The human CXCL8 ELISA was developed in our laboratory using monoclonal mouse anti-human CXCL8 (MAB208) and polyclonal goat anti-human CXCL8 (BAF208) antibodies from R&D Systems (6 (link)). Human CCL3 was measured with a specific ELISA Duoset kit as per the manufacturer’s instructions (R&D Systems).

Abouelasrar Salama S., De Bondt M., De Buck M., Berghmans N., Proost P., Oliveira V.L., Amaral F.A., Gouwy M., Van Damme J, & Struyf S. (2020). Serum Amyloid A1 (SAA1) Revisited: Restricted Leukocyte-Activating Properties of Homogeneous SAA1. Frontiers in Immunology, 11, 843.

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Inhibition of FAK and CXCL8 in RPE Cells

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An avirulent type 2 strain of T. gondii expressing green fluorescent protein (PTG strain, ATCC #50941) [13 (link)] was purchased from the American Type Culture Collection. Tachyzoites were maintained by serial passaging in HFF.
FAK was inhibited by treating 1 μM of PF-573228 (Tocris, Minneapolis, MN, USA) on apical side of RPE monolayer and CXCL8 was neutralized by treating 1 μg/mL of human CXCL8 antibody (MAB208; R&D systems, Minneapolis, MN, USA) on basolateral side of RPE monolayer. An antibody with corresponding IgG₁ isotype (MAB002; R&D systems) was used as a control.

Song H.B., Jun H.O., Kim J.H., Lee Y.H., Choi M.H, & Kim J.H. (2017). Disruption of outer blood-retinal barrier by Toxoplasma gondii-infected monocytes is mediated by paracrinely activated FAK signaling. PLoS ONE, 12(4), e0175159.

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Quantification of Chemokines by ELISA

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Quantification of chemokines in cell supernatants was done by ELISA. The human CXCL8 ELISA was developed in our laboratory using monoclonal mouse anti-human CXCL8 (MAB208) and biotinylated polyclonal goat anti-human CXCL8 (BAF208) antibodies purchased from R&D Systems (Minneapolis, MN, USA) [26 (link)]. Similarly, the human CCL2 ELISA was also developed in our laboratory with reagents from R&D Systems [monoclonal mouse anti-human CCL2 (MAB679) and biotinylated monoclonal mouse anti-human CCL2 (BAF279)] [28 (link)]. Human CCL3 and murine CCL2, CXCL1 and CXCL6 were measured with a specific Duoset ELISA kit following the manufacturer's instructions (R&D Systems).

Abouelasrar Salama S., De Bondt M., Berghmans N., Gouwy M., de Oliveira V.L., Oliveira S.C., Amaral F.A., Proost P., Van Damme J., Struyf S, & De Buck M. (2020). Biological Characterization of Commercial Recombinantly Expressed Immunomodulating Proteins Contaminated with Bacterial Products in the Year 2020: The SAA3 Case. Mediators of Inflammation, 2020, 6087109.

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Cell Migration Assay with IFNγ and IL-8

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The cell migration transwell assay was performed as described by the manufacturer’s protocol (Corning, NY, USA). SKOV3 cells were seeded onto the Corning Biocoat Matrigel Chambers (Corning #354480) at a density of 25,000 cells/0.5 mL in serum-free RPMI medium, with bottom wells containing RPMI medium with 10% FBS. Cells in the top chambers were incubated with recombinant human IFNγ (0 and 50 ng/mL; R&D Systems, 285-IF-100) in the presence of IL-8 neutralizing monoclonal antibody (2 μg/mL; R&D, MAB208) or control IgG (2 μg/mL; Santa Cruz Biotechnology; sc-2025) for 24 h. After incubation, non-migrating cells were scrubbed from the upper surface of the top chambers using cotton swabs. Migrating cells on the bottom membranes were fixed with 100% methanol, stained with crystal violet, washed, and air dried. Migrating cells were counted in five randomly selected fields under a phase-contrast microscope at 10X magnification, and quantified using ImageJ software as described [37 (link)].

Padmanabhan S., Gaire B., Zou Y., Uddin M.M., DeLeon D, & Vancurova I. (2021). IFNγ induces JAK1/STAT1/p65 NFκB-dependent interleukin-8 expression in ovarian cancer cells, resulting in their increased migration. The international journal of biochemistry & cell biology, 141, 106093.

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