The expression of IL-1β, IL-6, and TNF-α in lung tissue lysates was determined using the Bio-Plex Multiplex Immunoassay kit (Bio-Rad) according to the manufacturer’s instructions. All assays were performed at room temperature in 96-well round-bottomed microtiter plates protected from light. Measurements and data analyses were performed with the Bio-Plex system in combination with Bio-Plex Manager software.
Bio plex multiplex immunoassay kit
Manufactured by Bio-Rad
Sourced in Italy
The Bio-Plex Multiplex Immunoassay kit is a laboratory equipment product used for the simultaneous detection and quantification of multiple analytes in a single sample. It is designed to enable researchers to measure various proteins, cytokines, and other biomolecules efficiently and accurately.
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5 protocols using bio plex multiplex immunoassay kit
1
Multiplex Immunoassay for Inflammatory Cytokines
2
Cytokine Profiling of Murine Colitis
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Lamina propria mononuclear cells (LPMC) were obtained from mice with TNBS-induced colitis (6 days duration) and seeded into 96 well plates (200,000 cells/well) for stimulation with LPS (5 μg/ml) in RPMI complete medium for 36 h at 37 °C16 (link)17 (link). Alternatively, CD4+ cells were sorted from the MLN after 3–4 d duration of TNBS colitis and then seeded into 96 well plates (100,000 cells/well) for stimulation with plate-bound anti-CD3 (5 μg/ml) and soluble anti-CD28 (2 μg/ml) in RPMI complete medium for 72 h at 37 °C. After incubation, the culture supernatant was analyzed for cytokine content by ELISA. Measurement of cytokine/chemokine release in the DSS model was achieved using a Bio-Plex® Multiplex Immunoassay kit (BioRad, Italy).
3
Multiplex Immunoassay of Adipokines and Cytokines
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The expression of adipokines and cytokines (insulin, IL-1β, IL-6, TNF-α, and MCP-1) in serum and tissue lysates was determined using the Bio-Plex Multiplex Immunoassay kit (Bio-Rad), according to the manufacturer instructions. Blank values were subtracted from all readings. All assays were performed directly at room temperature in a 96-well round-bottomed microtiter plates (Muller Ratiolab, Dreieich, Germany) protected from light. Measurements and data analysis were performed with a Bio-Plex system in combination with Bio-Plex Manager software.
4
Tumor Growth Modulation by sEH Inhibitor
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1 × 106 LLc1 cells were subsequently injected in WT or KI male mice (12–14 weeks of age). For the study utilising genetically-modified LLc1 cells stably overexpressing WT or C521S sEH, 1 × 105 cells were injected in C57BL/6 mice. After the tumor size reached ~100 mm3, the mice were treated with vehicle (50% saline with 50% DMSO) or 5 mg/kg/day nitro-oleate or oleate using osmotic pumps (ALZET). Tumor volume, which was not allowed to exceed 1.5 cm3, was measured every 2 or 3 days using a calliper. sEH activity and protein expression (Cayman) were measured in harvested tumor tissues. In addition, paraffin-embedded sectioned tumor tissues were stained using Alexa Fluor™ 568 Conjugate isolectin B4 (Thermo Fisher scientific) and DAPI (Sigma). Stained sections were visualized using a Nikon eclipse TS100 fluorescence microscope and isolectin B4 positive cells were counted using Image J software [23 (link)]. Plasma was analysed for the EETs and DHETs as described below, and cytokines (interleukin (IL)-1β, IL-6, IL-10 and IL-12p70) analysed using a Bio-Plex multiplex immunoassay kit (Bio-Rad) with a Bio-Plex MAGPIX fluorescence magnetic bead-based immunoassay reader (Bio-Rad) according to the manufacturer's instructions.
5
LPS-Induced Organ Pathology and Inflammation
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