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Mkc00b

Manufactured by R&D Systems
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The MKC00B is a laboratory equipment product manufactured by R&D Systems. It is designed to perform a specific core function, but a detailed description while maintaining an unbiased and factual approach is not available.

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Lab products found in correlation

Mif00, by R&D Systems (2 mentions) Mta00b, by R&D Systems (2 mentions) Mm200, by R&D Systems (2 mentions) Mje00b, by R&D Systems (2 mentions) Dta00d, by R&D Systems (2 mentions) Mouse xl cytokine array, by R&D Systems (1 mentions) Fusion sl2 3500 wl, by Vilber (1 mentions) T per tissue protein extraction reagent, by Thermo Fisher Scientific (1 mentions) Tissuelyser 2, by Qiagen (1 mentions) Mx000, by R&D Systems (1 mentions) Mle 12 cells, by American Type Culture Collection (1 mentions) Mje00, by R&D Systems (1 mentions) Mcp 1, by R&D Systems (1 mentions) Dtga00, by R&D Systems (1 mentions) Mirin, by Selleck Chemicals (1 mentions) B2064, by Merck Group (1 mentions) β actin, by Santa Cruz Biotechnology (1 mentions) M1700, by R&D Systems (1 mentions) Mab3802, by Merck Group (1 mentions) Mgd150, by R&D Systems (1 mentions)

12 protocols using mkc00b

1

Cytokine profiling of ischemic colonic anastomosis

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Total protein lysates from ischemic colonic anastomotic tissue were prepared using T-PER Tissue Protein Extraction Reagent (Thermo Fisher Scientific) and Tissue Lyser II (Qiagen). Lysates were then analyzed with a Mouse XL Cytokine Array (ARY028, R&D Systems, Bio-Techne). Chemiluminescence was detected with Fusion SL2-3500.WL (Vilber Lourmat). Densitometric analysis of dots of each cytokine was performed with ImageJ. Background signal was subtracted, and expression of each cytokine was normalized to its own positive control.
In addition, IL-6 protein levels were determined in whole tissue lysates from septic colonic anastomoses using a commercially available ELISA (M6000B, R&D Systems, Bio-Techne). In CM from LPS-polarized proinflammatory (M1) BMDMs, IL-6 protein and CXCL-1 were also analyzed by ELISA (IL-6: M6000B, R&D Systems, Bio-Techne; CXCL-1: MKC00B, R&D Systems, Bio-Techne). Absorbance was measured at 450 nm. Further details on culture conditions are outlined below. All experiments were performed in triplicate.
Strowitzki M.J., Kimmer G., Wehrmann J., Ritter A.S., Radhakrishnan P., Opitz V.M., Tuffs C., Biller M., Kugler J., Keppler U., Harnoss J.M., Klose J., Schmidt T., Blanco A., Taylor C.T, & Schneider M. (2021). Inhibition of HIF-prolyl hydroxylases improves healing of intestinal anastomoses. JCI Insight, 6(8), e139191.
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2

Chemotaxis Assay for Neutrophils

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Boyden chambers of 3 µm pore size (Corning Incorporated, Corning, NY, USA) was used to assess chemotaxis for neutrophils. MLE-12 cells (mouse lung epithelial cell lines, ATCC CRL-2110; American Type Culture Collection, Manassas, VA, USA) were plated in 24-well plates (4×105 per well) and incubated with the three groups of lung CD4+ T cells (blank control group, negative control group and SOCS3 overexpressing group) for 48 h. The culture supernatant was harvested. Then, the treated supernatant was added to the lower chambers, while neutrophils were added to the top chambers for incubation for another 90 min at 37°C in a humidified atmosphere with 5% CO2. The filters were fixed with ethanol and stained with DAPI to visualize the nuclei. The number of cells that had migrated through the entire thickness of the filter was evaluated, and the results were expressed as the chemotactic index, being the number of cells that migrated towards the sample divided by the number of cells that migrated towards the control medium. Triplicate chambers were used in each experiment and five fields were examined in each filter. CXCL1 and CXCL5 were measured in the culture supernatant using commercially available ELISA kits (catalog nos. MKC00B and MX000; R&D Systems, Inc., Minneapolis, MN, USA) according to the manufacturer's protocol.
Ding F.M., Liao R.M., Chen Y.Q., Xie G.G., Zhang P.Y., Shao P, & Zhang M. (2017). Upregulation of SOCS3 in lung CD4+ T cells in a mouse model of chronic PA lung infection and suppression of Th17-mediated neutrophil recruitment in exogenous SOCS3 transfer in vitro. Molecular Medicine Reports, 16(1), 778-786.
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3

Cytokine quantification in BALF

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The concentration of CXCL1, CXCL2, IL6 and IFNg in BALF supernatants were measured by ELISA kits following the manufacturer’s protocol. ELISA kits for mouse CXCL1 (MKC00B), mouse CXCL2 (MM200), mouse IL6 (M6000B) and mouse IFNg (MIF00) were purchased from R&D systems (Minneapolis, MN, USA).
Huang H.Q., Li N., Li D.Y., Jing D., Liu Z.Y., Xu X.C., Chen H.P., Dong L.L., Zhang M., Ying S.M., Li W., Shen H.H., Li Z.Y, & Chen Z.H. (2021). Autophagy Promotes Cigarette Smoke-Initiated and Elastin-Driven Bronchitis-Like Airway Inflammation in Mice. Frontiers in Immunology, 12, 594330.
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4

Cytokine Quantification in Cell and Tissue

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Levels of IL-6, TNF-α, CXCL-1 and MCP-1 in cell culture supernatants, serum and heart tissue (10% wt/vol in ice cold PBS buffer) were determined using commercially available ELISA kits following manufacture’s specifications (IL-6, M6000B; TNF-α, MTA00B; CXCL-1, MKC00B; MCP-1, MJE00; R&D Systems).
Qin X., Peterson M.R., Haller S.E., Cao L., Thomas D.P, & He G. (2018). Caspase recruitment domain-containing protein 9 (CARD9) knockout reduces regional ischemia/reperfusion injury through an attenuated inflammatory response. PLoS ONE, 13(6), e0199711.
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5

BALF Cytokine Quantification by ELISA

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The supernatant fraction of the BALF was thawed on ice and used in enzyme-linked immunosorbant assays (ELISA). The mouse homologs of human interleukin (IL)-8, Keratinocyte-Derived Cytokine (KC) and Macrophage Inflammatory Protein (MIP)-2, were detected using ELISA (R&D Systems, Product Number MKC00B and MM200, respectively). TGF-α was also analyzed similarly (R&D Systems, Product number DTGA00). BALF cytokine concentrations were determined by comparison to standard curves for each cytokine provided by the supplier.
Chang S., Linderholm A, & Harper R. (2015). DUOX-Mediated Signaling Is Not Required for LPS-Induced Neutrophilic Response in the Airways. PLoS ONE, 10(7), e0131810.
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6

Antibody-based Protein Analysis in DNA Damage

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Antibodies against CC10 (Santa Cruz Biotechnology, sc-9772), CD68(Servicebio, GB11067), Actin B (Santa Cruz Biotechnology, sc-47778), RAD50(Abcam, ab89), γH2AX (EMD Millipore, 05-636), RPA (Abcam, ab2175), pATM (EMD Millipore) and 53BP1 (EMD Millipore, MAB3802) were used. Antibodies for immunofluorescence were diluted at 1:1000 in 5% BSA (Sigma-Aldrich, B2064) except for the pATM which was diluted at 1:500. Antibodies for western blot were diluted at 1:1000 in 5% nonfat dried milk. All primers used in the study were synthesized by Sangon Biotech,Shanghai. ELISA kits for mouse Cxcl1 (MKC00B), mouse Cxcl2 (MM200), mouse Il6 (M6000B), mouse Il17(R&D, M1700) and mouse Ifn-γ (R&D, MIF00) were purchased from R&D systems. MRE11 inhibitor mirin was purchased from Selleck (S8096), and its final concentration in culture medium was 100μM.
Luo M., Bao Z., Xu F., Wang X., Li F., Li W., Chen Z., Ying S, & Shen H. (2018). Unrepaired DNA damage in macrophages causes elevation of particulate matter- induced airway inflammatory response. Aging (Albany NY), 10(4), 549-560.
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7

CXCL1 Quantification in Cell Supernatants

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Supernatants from treated cells were collected. Cell debris was removed by centrifugation at 6,000 rpm for 1 min. Samples were diluted with the dilution buffer provided in each ELISA kit, including murine CXCL1 (MKC00B, R&D). All experiments were performed in triplicate or more.
Wang W., Prokopec J.S., Zhang Y., Sukhoplyasova M., Shinglot H., Wang M.T., Linkermann A., Stewart-Ornstein J, & Gong Y.N. (2022). Sensing plasma membrane pore formation induces chemokine production in survivors of regulated necrosis. Developmental cell, 57(2), 228-245.e6.
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8

Measuring Growth Factors and Insulin

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GDF15 and mouse KCGRO were measured using kits from R&D systems (#MGD150 and #MKC00B), and insulin was measured using the Alpco kit (#80-INSMS-E01). Rat KCGRO was measured using the MSD Kit (#K15059D). The protocols provided with the kits were followed.
Breen D.M., Jagarlapudi S., Patel A., Zou C., Joaquim S., Li X., Kang L., Pang J., Hales K., Ziso-Qejvanaj E., Vera N.B., Bennett D., He T., Lambert M., Kelleher K., Wu Z., Zhang B.B., Lin L., Seeley R.J, & Bezy O. (2021). Growth differentiation factor 15 neutralization does not impact anorexia or survival in lipopolysaccharide-induced inflammation. iScience, 24(6), 102554.
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9

Quantifying Inflammatory Cytokines in Mice

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The concentrations of IL-6, IL-1β, CXCL2, CXCL1 or CCL2 in the peritoneal lavage fluid, plasma, or bone marrow of mice were determined with ELISA kits (R&D Systems, M6000B, MLB00C, MM200, MKC00B, MJE00B) according to the manufacturer’s instructions.
Liu Y., Song R., Zhao L., Lu Z., Li Y., Zhan X., Lu F., Yang J., Niu Y, & Cao X. (2022). m6A demethylase ALKBH5 is required for antibacterial innate defense by intrinsic motivation of neutrophil migration. Signal Transduction and Targeted Therapy, 7, 194.
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10

Profiling Cytokine Levels in Cell Culture and Mouse Models

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The concentrations of interleukin (IL)‐6, IL‐8, IL‐1β, and TNFα proteins in cell culture supernatants and mouse cytokines such as IL‐6 and CXCL122, 23 from bronchoalveolar lavage fluid (BALF) supernatants and circulating plasma were measured by enzyme‐linked immunosorbent assay (ELISA) kits under the manufacturer's protocol. These ELISA kits were human IL‐6 (D6050; R&D Systems), human IL‐8 (D8000C; R&D Systems), human IL‐1β (DLB50; R&D Systems), human TNFα (DTA00D; R&D Systems), mouse IL‐6 (M6000B; R&D Systems), and mouse CXCL1 (MKC00B; R&D Systems).
Fu H., Liu X., Shi L., Wang L., Fang H., Wang X, & Song D. (2023). Regulatory roles of Osteopontin in lung epithelial inflammation and epithelial‐telocyte interaction. Clinical and Translational Medicine, 13(8), e1381.
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