Msd mouse proinflammatory 5 plex plus kit

Manufactured by Mesoscale

Sourced in United States

The MSD Mouse Proinflammatory V-Plex Plus Kit is a multiplex assay designed to quantitatively measure multiple analytes in mouse biological samples. The kit utilizes electrochemiluminescence (ECL) technology to detect and quantify the levels of various inflammatory markers simultaneously.

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Lab products found in correlation

Sector imager 6000 plate reader, by Mesoscale (2 mentions) Micro bca protein assay kit, by Thermo Fisher Scientific (1 mentions) Mouse tnf ri and tnfr 2 ultra sensitive kits, by Mesoscale (1 mentions) Sector imager 6000, by Mesoscale (1 mentions) Quickplex sq 120, by Mesoscale (1 mentions) Phosphatase inhibitor cocktail 1, by Merck Group (1 mentions) Complete protease inhibitor cocktail, by Roche (1 mentions) Mouse tlr2 elisa kit, by Abcam (1 mentions) Msd quickplex sq120 plate reader, by Mesoscale (1 mentions) Phosphatase inhibitor, by Merck Group (1 mentions) U plex kit, by Mesoscale (1 mentions)

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MSD V-Plex (3 citations) Mouse Proinflammatory V-Plex Plus Kit (2 citations) MSD kits (2 citations)

5 protocols using msd mouse proinflammatory 5 plex plus kit

Measuring Brain Cytokines and Receptors

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One series of brain tissue was lysed in Complete Mesoscale buffer according to the manufacturer’s protocol (Mesoscale Discovery, Rockville, MD, United States). Protein concentrations were determined according to the protocol from Micro BCA protein assay kit (Thermo Fischer Scientific). Cytokine and chemokine concentrations were estimated using an MSD Mouse Pro-Inflammatory V-Plex Plus Kit (Mesoscale Discovery) and TNF receptor concentrations using the Mouse TNF-RI and TNFR-II Ultra-Sensitive Kits (Mesoscale Discovery). XPro1595 levels were measured in ischemic brain tissue lysates using a human TNF V-Plex immunoassay (Mesoscale Discovery) as previously described (Karamita et al., 2017 (link)). The standard in the kit was replaced with XPro1595, which was diluted in the kit diluent number 2. Etanercept levels were measured using a human TNFRII Ultra-Sensitive immunoassay (Mesoscale Discovery) (Karamita et al., 2017 (link)). The standard in the kit was replaced with etanercept diluted in the kit diluent number 2. All kits were read on a SECTOR Imager 6000 plate reader (Mesoscale Discovery) according to the manufacturer’s instructions. All samples were run in duplex, and coefficient of variation (CV) values below 25% were accepted.

Yli-Karjanmaa M., Clausen B.H., Degn M., Novrup H.G., Ellman D.G., Toft-Jensen P., Szymkowski D.E., Stensballe A., Meyer M., Brambilla R, & Lambertsen K.L. (2019). Topical Administration of a Soluble TNF Inhibitor Reduces Infarct Volume After Focal Cerebral Ischemia in Mice. Frontiers in Neuroscience, 13, 781.

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Cytokine Quantification Post-SCI

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To measure cytokine protein levels by the MSD Mouse Proinflammatory V-Plex Plus Kit (IFNγ, IL-1β, IL-2, IL-4, IL-5, IL-6, IL-10, IL-12p70, CXCL1, TNF; K15012C, Mesoscale) under naïve conditions and 3 days after SCI, we used a SECTOR Imager 6000 (Mesoscale Discovery, Rockville, USA) Plate Reader according to the manufacturer’s instructions. The same samples as those used for Western blotting were diluted two-fold in Diluent 41 prior to measurement and 50 µl dilution was loaded in each well. Data was analyzed using MSD Discovery Workbench software [50 (link), 51 (link)].

Ellman D.G., Isaksen T.J., Lund M.C., Dursun S., Wirenfeldt M., Jørgensen L.H., Lykke-Hartmann K, & Lambertsen K.L. (2017). The loss-of-function disease-mutation G301R in the Na+/K+-ATPase α2 isoform decreases lesion volume and improves functional outcome after acute spinal cord injury in mice. BMC Neuroscience, 18, 66.

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Multiplex Cytokine Quantification in Hippocampus

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The MSD Mouse Proinflammatory V-Plex Plus kit (INF-γ, IL-1β, IL-12p70, IL-2, IL-4, IL-5, IL-6 IL-10, CXCL1, TNF-α; K15012C, Mesoscale) was used to measure the concentrations of different cytokines in the pooled S1 and S2 fractions of homogenized hippocampus (25 µl/sample) by using a QuickPlex SQ120 (Mesoscale Discovery, Rockville, USA) Plate Reader according to the manufacturer’s instructions and as described previously^{46 (link)}. The MSD Discovery Workbench software was used to analyze the data. The concentrations were normalized to protein concentrations measured with the Bradford assay.

Svensson M., Olsson G., Yang Y., Bachiller S., Ekemohn M., Ekstrand J, & Deierborg T. (2021). The effect of electroconvulsive therapy on neuroinflammation, behavior and amyloid plaques in the 5xFAD mouse model of Alzheimer’s disease. Scientific Reports, 11, 4910.

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Multiplex Cytokine Analysis in CNS Samples

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Multiplex analysis was performed in spinal cord tissues (thoracic segments), and in OPC and OL cultures. Spinal cord samples were obtained from the thoracic spinal cord of PBS-perfused mice homogenized in 300 μl RIPA buffer containing phosphatase inhibitor cocktail 1 (Sigma) and Complete protease inhibitor cocktail (Roche Diagnostics). OPC and OL samples were obtained by homogenizing cells obtained from 2 wells of a 6-well plate in 250 μl RIPA buffer with phosphatase and protease inhibitors. Cytokine expression was evaluated as previously described (Madsen et al., 2015 ) using the MSD Mouse Proinflammatory V-Plex Plus Kit, or individual plates for the assessment of CCL2, CCL5 and TNFR1 (Mesoscale). Samples were run on a SECTOR Imager 6000 Plate Reader (Mesoscale Discovery) and the data analyzed with the MSD Discovery Workbench software. Samples with CV values above 25% were excluded.

Madsen P.M., Desu H.L., Vaccari J.P., Florimon Y., Ellman D.G., Keane R.W., Clausen B.H., Lambertsen K.L, & Brambilla R. (2019). Oligodendrocytes modulate the immune-inflammatory response in EAE via TNFR2 signaling. Brain, behavior, and immunity, 84, 132-146.

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Protein Quantification in Brain Tissue

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For protein determination, one series of brain tissue was sonicated in cold phosphate-buffered saline (PBS) (Sigma life science, St. Louis, MO, USA) containing phosphatase inhibitors (Sigma-Aldrich, Soeborg, DK) and cOmplete mini, EDTA-free proteinase inhibitor cocktail (Roche, Basel, Switzerland). The total protein content was measured using the bicinchoninic acid (BCA) assay according to the manufacturer’s protocol (P4417-100TAB, Thermo Scientific, Waltham, MA, USA).
Electrochemiluminescence analysis was performed on brain, serum, cells, and media using the MSD Mouse Proinflammatory V-Plex Plus Kit (IFNγ, IL-2, IL-4, IL-5, IL-6, IL-10, IL-12p70, CXCL1, TNF; K15012C, Mesoscale Discovery), the U-Plex Kit (TNFR1; K15069L, Mesoscale), and the R-Plex kit (TNFR2; K150ZSR-2, Mesoscale Discovery) according to the manufacturer’s instructions. Plates were read using the MSD QuickPlex (SQ120) Plate Reader (Mesoscale Discovery), and the data analyzed using the MSD Discovery Workbench software as detailed [20 (link)].
IL-1Ra ELISA was performed as specified in the Quantikine mouse IL-1Ra assay (MRA00, R&D). TLR2 ELISA was performed as specified in the mouse TLR2 Elisa kit (ab224880, Abcam). The measured protein was normalized to the total protein content measured using the BCA assay [32 (link)].

von Linstow C.U., Hindkjær S.M., Nielsen P.V., Degn M., Lambertsen K.L., Finsen B, & Clausen B.H. (2021). Bone Marrow-Derived IL-1Ra Increases TNF Levels Poststroke. Cells, 10(4), 956.

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