Bio-Plex Pro mouse cytokine 23-plex assay was performed to test for concentrations of 23 cytokines in MDSC-CM or Control-CM. Three MDSC-CM samples and two control-CM samples, generated in independent experiments, were assayed. The cytokines assessed in CM were: IL-1α, IL-1β, IL-2, IL-3, IL-4, IL-5, IL-6, IL-9, IL-10, IL-12(p40), IL-12(p70), IL-13, IL-17A, G-CSF, GM-CSF, IFN-γ, KC, MCP, MIP-1α, MIP-1β, RANTES, eotaxin, TNF-α. Cytokines not shown in Fig. 4 did not show differences between MDSC-CM and control-CM.
Pro mouse cytokine 23 plex assay
Manufactured by Bio-Rad
The Pro Mouse Cytokine 23-plex Assay is a multiplexed immunoassay designed to detect and quantify the levels of 23 different mouse cytokines simultaneously in a single sample. The assay utilizes magnetic beads coated with specific antibodies to capture the target cytokines, which are then detected and quantified using a flow-based detection system.
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Lab products found in correlation
Cell lysis kit, by Bio-Rad (2 mentions)
Bio plex, by Bio-Rad (1 mentions)
Sample diluent, by Bio-Rad (1 mentions)
Facscanto 2, by BD (1 mentions)
Cytometric bead array cba mouse th1 th2 th17 cytokine kit, by BD (1 mentions)
Fcap array v3, by BD (1 mentions)
Luminex 200 system, by Bio-Rad (1 mentions)
8 protocols using pro mouse cytokine 23 plex assay
1
Cytokine profiling of MDSC conditioned media
2
Cytokine Profiling of Influenza-Infected Macrophages
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A549 cells were infected with either X31 (H3N2) or NL/09 (H1N1) at a multiplicity of infection (MOI) of 3 for 18 h in the absence of TPCK-treated trypsin. MAb 2B9, FEE8, 9H10, 6F12, or control IgG (anti-GST mAb) were added at a concentration of 50 µg per mL in 50 µL. Alveolar macrophages from freshly collected BALF of naive mice were subsequently added at a concentration of 1000 cells in 50 µL. The ratio of virus-infected cells to primary alveolar macrophages was 4:1. The FcγR block was a mixture of anti-CD32 and anti-CD16 antibodies (BD Biosciences; cat. no. 553141) (1 µg per million cells in 100 µL). Seventy-two hours later, cell culture supernatants were collected and assessed for cytokine production using a Bio-Plex Pro Mouse Cytokine 23-plex assay as described above. Fold induction was normalized to IgG control and a heat map was generated using GraphPad Prism.
3
Cytokine Profiling from Tissue
4
Cytokine Profiling from Tissue
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Proteins were extracted from 25 mg of tissue using Bio-Plex cell lysis kit. Cytokine concentrations were measured using Bio-Plex Pro Mouse Cytokine 23-plex Assay according to manufacturers protocol.
5
Cytokine Profile in Opa1-/- Mice
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After 90 days of treatment, plasma was obtained from blood collected from untreated Opa1–/– mice and mice treated with the individual compounds Met, Gal, and the combination drug RJx-01. The blood cytokines panel (Bio-Plex Pro Mouse Cytokine 23-plex Assay #M60009RDPD) was measured following the manufacturer’s instructions.
6
Multiplex Cytokine Profiling
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Serum cytokines were assayed by a Bio-Plex Pro Mouse Cytokine 23-plex Assay (M60009RDPD) using Luminex Technology according to the manufacturer’s protocol (Bio-Plex, Bio-Rad Laboratories).
7
Inflammatory Cytokine Profiling in Pregnant Mice
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Bio-Plex Pro Mouse Cytokine 23-plex assay (Bio-Rad, M60009RDPD) was used to compare the concentration of inflammatory cytokines and chemokines in hippocampal cytosolic fractions from ASIC2a +/+ and +/- pregnant mice. Samples were thawed and diluted 1:2 with Bio-Plex sample diluent and run following manufacturer’s directions. Standard curves were generated for each analyte and sample concentrations determined. All measured concentrations were normalized to protein concentration in each sample.
8
Pneumonia Cytokine Response Measurement
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Anesthetize the mice with 2,2,2-tribromoethanol (3.75–5 mg/25 g), then slowly pipet the inoculum with 2 × 108 CFU into the nostrils of the mice. The orbital venous plexus blood of mice was collected 48 h after infection, and the BALF was collected after euthanizing the mice. The level of mouse plasma cytokine was detected by Bio-Plex Pro Mouse Cytokine 23-plex Assay in accordance with the manufacturer’s instructions (Bio-Rad, Luminex 200™ System). Cytokine levels in mouse BALF were detected by BD Cytometric Bead Array (CBA) Mouse Th1/Th2/Th17 Cytokine Kit. BD FACSCantoII was used for data collection and FCAP Array V3.0 was used for data analysis. IL-1β in the BALF and mouse BMDMs supernatants was measured by using ELISA kit (R&D systems). IL-1β and tumor necrosis factor α (TNF-α) released by THP-1 cells were detected by Human IL-1β or TNF-α ELISA kit (Sangon Biotech), and interleukin-17A (IL-17A) released by THP-1 was measured by Human IL-17 Quantikine HS ELISA kit (R&D systems).
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