HEK 293A transfected cells were stimulated for 20 h with 0.5, 5, or 50 μg/ml of RA , RAinc , K98 , and K98inc or 100 ng/ml LPS or 1 nM P3C or 10 ng/ml FSL-1 or 0.5% EtOH (vehicle) as control. IL-8 levels were determined in cell supernatants by ELISA using Human CXCL8/IL-8 DuoSet ELISA Kit (R&D Systems, Minneapolis, MN, USA).
Human il 8 cxcl8 duoset elisa kit
Manufactured by R&D Systems
Sourced in United States
The Human IL-8/CXCL8 DuoSet ELISA kit is a reagent set used for the quantitative determination of human interleukin-8/CXCL8 levels in cell culture supernatants, serum, and plasma samples. The kit includes the necessary components to perform an enzyme-linked immunosorbent assay (ELISA).
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Rneasy plus kit, by Qiagen (1 mentions)
Human ccl2 mcp 1 quantikine elisa kit, by R&D Systems (1 mentions)
Microplate spectrophotometer, by Agilent Technologies (1 mentions)
Hyaluronan duoset elisa kit, by R&D Systems (1 mentions)
Luciferase assay kit, by Promega (1 mentions)
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Human myeloperoxidase duoset elisa kit, by R&D Systems (1 mentions)
21 protocols using human il 8 cxcl8 duoset elisa kit
1
HEK 293A Inflammatory Response Assay
2
Microfluidic Culture System for CXCL8/IL-8 Monitoring
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In the microfluidic culture system the basolateral flow was collected at 2hr intervals with the automated fraction collector. In static control experiments basolateral supernatant was collected at matching time points. The concentration of CXCL8/IL–8 in supernatants was analysed by Human CXCL8/IL–8 DuoSet ELISA kit (R&D Systems, Abingdon, UK). The physical barrier properties before and after stimulation were monitored by measuring TER.
3
Evaluating TNFα-Induced IL-8 Modulation
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HCT116 (ATCC CCL-247) colon cell line was grown at 37 • C in a humidified 5% CO 2 -95% air atmosphere. Cells were maintained in McCoy s 5a Modified Medium. Cells were seeded, in triplicate, into a 24-well plate at a concentration of 50,000 cells per well in 500µL of growing media, and then incubated for 24 h at 37 • C in a humidified 5% CO 2 -95% air atmosphere. When cell excitation was performed with TNFα, cultures in each well were treated with 50 ng/mL recombinant human TNFα (PeproTech, Rocky Hill, NJ, USA) with or without HSB tuber extract. The supernatant was taken and the levels of IL-8 were measured per 1 g of dry weight of HSB tuber crude extract at 16 h post-treatment using the commercial Human CXCL8/IL-8 DuoSet ELISA kit (R&D Systems, Minneapolis, MN, USA). In each in-vitro experiment, a Tukey-Kramer HSD test (p ≤ 0.05; n = 3) was executed. The results were normalized to the TNFα control and expressed in relative units (%).
4
Plasma CXCL8 Quantification by ELISA
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CXCL8 was measured in plasma using the R&D Systems DuoSet Human IL-8/CXCL8 ELISA Kit (catalogue No. DY208) as per manufacturer’s instructions.
5
PYO-induced IL-8 Secretion Quantification
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The confluent CFBE Tet-on cells were treated with 1 µg/mL Dox for 2 days, followed by a 1-day treatment with 1 µg/mL Dox and 5 µM PYO for 24 h at 37 °C. After the PYO treatment, the culture medium was collected and used for ELISA after centrifugation at 12,000 rpm for 5 min. A fresh cell culture medium was used to measure the background signal. Human IL-8 was measured in cell culture media by the DuoSet® Human IL-8/CXCL8 ELISA kit (R&D Systems, Minneapolis, MN, USA) according to the manufacturer’s instructions. PYO-induced IL-8 secretion was quantified by subtracting the IL-8 secretion in DMSO-treated cells from that in PYO-treated cells.
6
Quantification of IL-8 Expression
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Total RNA was isolated using the RNeasy plus Kit (Qiagen) and reverse-transcribed to cDNA according to the manufacturer’s protocols (MultiScribe Reverse Transcriptase; Applied Biosystems). Relative transcriptional level were measured by SYBR Green dye-based quantitative real-time PCR (qRT–PCR) and analyzed using the ABI Prism 7500 Fast RT-PCR System (Applied Biosystems). GAPDH was used as a housekeeping marker. The list of primers is given in Supplementary Table 1 . IL-8 secretion in the supernatant was quatified using the Human IL-8/CXCL8 DuoSet ELISA kit (R&D) and measured using a Synergy H1 microplate reader (BioTek Instruments) according to the manufacturer’s protocol.
7
Quantifying IL-8 in Cell Culture
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Measurements of the quantity of IL-8 in the cell culture medium was performed using Human IL-8/CXCL8 DuoSet ELISA kit from R&D systems according to manufacturer’s instructions.
8
Quantifying Interleukin-8 in GBM
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The levels of Interleukin-8 in the GBM supernatants were assessed with the Human IL-8/CXCL8 DuoSet ELISA kit according to the protocol provided by the manufacturer (R&D Systems, Minneapolis, MN, USA). Absorbance was assessed at 450–540 nm with a TECAN plate reader. The analysis was performed independently for different batches of supernatants.
9
Quantifying Human IL-8 Levels via ELISA
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IL-8 ELISA was performed using the human IL-8/CXCL8 DuoSet ELISA kit (R&D Systems) according to the manufacturer’s instructions.
10
Quantification of IL-8 and MCP-1 in hCASMCs
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Enzyme-linked immunosorbent assays (ELISAs) were performed to measure IL-8 and MCP-1 protein levels in lysates of hCASMC treated with LPS. To obtain cell lysates, cells were harvested in cold PBS and sonicated 3 × 10 s on ice. The lysate was then centrifuged at 1800 × g for 5 min at 4°C and the supernatant was collected. The assays were performed using the Human IL-8/CXCL8 DuoSet ELISA kit (#DY208) and the Human CCL2/MCP-1 Quantikine ELISA kit (#DCP00), both from R&D Systems. We adhered to protocols provided by the manufacturer.
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