Mouse il 1 beta il 1f2 duoset elisa kit

Manufactured by R&D Systems

Sourced in United States, France

The Mouse IL-1 beta/IL-1F2 DuoSet ELISA kit is a quantitative sandwich enzyme-linked immunosorbent assay (ELISA) for the measurement of mouse interleukin-1 beta (IL-1 beta) and interleukin-1 family member 2 (IL-1F2) in cell culture supernates, cell lysates, tissue homogenates, and serum.

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8 protocols using mouse il 1 beta il 1f2 duoset elisa kit

Cytokine Quantification in Murine Infection

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Blood samples were collected at different days of infection and the sera were stored at - 80°C. Samples from individual mice were diluted 10x for IL-1β quantification or non-diluted, for IL-18 detection. Cytokine levels were determined using mouse IL-18/IL-1F4 ELISA kit and mouse IL-1beta/IL-1F2 DuoSet ELISA kit (R and D Systems) according to the manufacturer's instructions.

Oliveira A.C., Gomes-Neto J.F., Barbosa C.H., Granato A., Reis B.S., Santos B.M., Fucs R., Canto F.B., Nakaya H.I., Nóbrega A, & Bellio M. (2017). Crucial role for T cell-intrinsic IL-18R-MyD88 signaling in cognate immune response to intracellular parasite infection. eLife, 6, e30883.

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Cytokine Expression Analysis in Mice

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For investigation of the effect of FDP on the expression of cytokines, a subset of mice were sacrificed and brain regions and plasma were harvested. Brain regions were lysed with 1X Cell Lysis Buffer supplemented with 1 mM PMSF (Cell Signaling, Danvers MA) and Protease Inhibitor Cocktail (Sigma-Aldrich, cat no. 11873580001). Brain tissue IL-1β was measured with Mouse IL-1 beta/IL-1F2 DuoSet ELISA Kit (R&D Systems, Minneapolis MN) according to the manufacturer’s instructions. For mouse primary microglia and human THP-1 macrophages, tissue culture supernatant was harvested for analysis of secreted cytokines. In human THP-1 cells, IL-1β was measured with Human IL-1 beta/IL-1F2 DuoSet ELISA Kit (R&D Systems) according to the manufacturer’s instructions.

Smith C., Trageser K.J., Wu H., Herman F.J., Iqbal U.H., Sebastian-Valverde M., Frolinger T., Zeng E, & Pasinetti G.M. (2021). Anxiolytic effects of NLRP3 inflammasome inhibition in a model of chronic sleep deprivation. Translational Psychiatry, 11, 52.

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Cytokine Quantification in Mice

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The IL-1β levels were determined using a mouse IL-1 beta/IL-1F2 DuoSet ELISA kit (R&D Systems, Minneapolis, MN, USA), and TNF-α levels were measured using a mouse TNF-α ELISA Ready-SET-Go kit (eBioscience, San Diego, CA, USA), according to the manufacturer's instruction. The absorbance measurements were read on a Multiskan GO microplate spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA).

Kwak S.B., Koppula S., In E.J., Sun X., Kim Y.K., Kim M.K., Lee K.H, & Kang T.B. (2018). Artemisia Extract Suppresses NLRP3 and AIM2 Inflammasome Activation by Inhibition of ASC Phosphorylation. Mediators of Inflammation, 2018, 6054069.

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Multiplex Cytokine Quantification

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Secreted human IL-1β and IL-10 were measured using the Human IL-1β/IL-1F2 DuoSet Kit or the Human IL-10 Quantikine Elisa kit (R&D Systems, DY201 and D1000B, respectively). Additional cytokines, growth factors and proteins (TNF, G-CSF, IL-6, IL-5, FGF, Arginase, IL-1RA, etc.) were quantified using a custom-assembled Human Multiplex Cytokine Assay (Bio-Plex; Bio-Rad Laboratories Inc.). Mouse IL-1β measured by either utilizing the Mouse IL-1 beta/IL-1F2 DuoSet ELISA kit (R&D Systems, DY401; recognizes predominantly mature IL-1β), via intracellular staining, or with the LEGENDplex^™ Mouse Inflammation Panel (13-plex) (BioLegend, 140150). The latter kit was also used to assess the additional mouse cytokines measured in this study.

Niyonzima N., Rahman J., Kunz N., West E.E., Freiwald T., Desai J.V., Merle N.S., Gidon A., Sporsheim B., Lionakis M.S., Evensen K., Lindberg B., Skagen K., Skjelland M., Singh P., Haug M., Ruseva M.M., Kolev M., Bibby J., Marshall O., O’Brien B., Deeks N., Afzali B., Clark R.J., Woodruff T.M., Pryor M., Yang Z.H., Remaley A.T., Mollnes T.E., Hewitt S.M., Yan B., Kazemian M., Kiss M.G., Binder C.J., Halvorsen B., Espevik T, & Kemper C. (2021). Mitochondrial C5aR1 activity in macrophages controls IL-1β production underlying sterile inflammation. Science immunology, 6(66), eabf2489.

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Colon Cytokine Quantification Protocol

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A 0.5 cm-long colon segment was collected about 1 cm from the rectum from each well-flushed mouse colon. These colon segments were cultured in 24-well plates containing 0.6 mL of complete tissue culture medium (DMEM with 25 mM HEPES, 0.05 mM 2-mercaptoethanol, 2 mM L-Glutamine, 100 U/mL Penicillin, 100 μg/mL Streptomycin, and 10% FBS) for 24 h till cell-free culture supernatant was collected. The collected supernatants were then subject to quantification of cytokine levels using the following ELISA kits: Mouse IL-1 beta/IL-1F2 DuoSet ELISA kit (#DY401-05), Mouse IL-6 DuoSet ELISA (#DY406-05), Mouse IL-10 DuoSet ELISA (#DY417-05), Mouse CXCL1/KC DuoSet ELISA (#DY453-05), Mouse Serum Amyloid A DuoSet ELISA (#DY2948-05), Mouse Cytokine Antibody Array, Panel A (#ARY006) (from R&D Systems, Minneapolis, MN), and Mouse SAA-3 ELISA (#EZMSAA3-12K) (from Millipore Sigma, Burlington, MA).

Lin N., Shay J.E., Xie H., Lee D.S., Skuli N., Tang Q., Zhou Z., Azzam A., Meng H., Wang H., FitzGerald G.A, & Simon M.C. (2018). Myeloid Cell Hypoxia-Inducible Factors Promote Resolution of Inflammation in Experimental Colitis. Frontiers in Immunology, 9, 2565.

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Multiplex ELISA Quantification of Inflammatory Cytokines

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IL-1β in cell culture supernatants and brain tissue samples—after the different treatment groups—was quantified using the mouse IL-1 beta/IL-1F2 DuoSet ELISA kit (R&D systems), IL-6 in the cell culture supernatant was measured through the mouse IL-6 ELISA kit (Thermo Fisher), TNF-α in the cell culture supernatants was determined using the mouse TNF-alpha DuoSet ELISA (R&D systems), and IL-18 in brain tissue was measured through the IL-18 mouse ELISA Kit (Invitrogen) following the manufacturers' recommended protocol. The IL-1β precursor and cleaved IL-1β in the FC of mice treated with vehicle, LPS, or C77-LPS were assessed by Western blot. All samples were run in technical duplicates.

Sebastian-Valverde M., Wu H., Al Rahim M., Sanchez R., Kumar K., De Vita R.J, & Pasinetti G.M. (2021). Discovery and characterization of small-molecule inhibitors of NLRP3 and NLRC4 inflammasomes. The Journal of Biological Chemistry, 296, 100597.

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Multiplex Cytokine Profiling Protocol

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Cell-free supernatants from in vitro cultures or tissue homogenates were used to detect cytokines by ELISA or Luminex analysis using a 32 plex MILLIPLEX MAP Mouse Cytokine/Chemokine Magnetic Bead Panel (Millipore), following the manufacturer's instructions. IL-1β levels were assayed using a Mouse IL-1beta/IL1F2 DuoSet ELISA kit (R&D Systems). IL-18 levels were measured by Mouse IL-18 ELISA (MBL), as previously described (44 (link)). Caspase-1 levels were determined in the supernatants using a Caspase-1 ELISA Kit (AdipoGen).

Javanmard Khameneh H., Leong K.W., Mencarelli A., Vacca M., Mambwe B., Neo K., Tay A., Zolezzi F., Lee B, & Mortellaro A. (2019). The Inflammasome Adaptor ASC Intrinsically Limits CD4+ T-Cell Proliferation to Help Maintain Intestinal Homeostasis. Frontiers in Immunology, 10, 1566.

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Quantification of IL-1β in MSC-2 and Plasma

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IL-1β concentration was measured using Mouse IL-1beta/IL-1F2 DuoSet ELISA Kit (R&D Systems, France) according to manufacturer’s instructions. MSC-2 culture media was collected 24 h after treatment and plasma was sampled 48 h after 5-FU injection in tumor-bearing mice.

Dumont A., de Rosny C., Kieu T.L., Perrey S., Berger H., Fluckiger A., Muller T., Pais de Barros J.P., Pichon L., Hichami A., Thomas C., Rébé C., Ghiringhelli F, & Rialland M. (2019). Docosahexaenoic acid inhibits both NLRP3 inflammasome assembly and JNK-mediated mature IL-1β secretion in 5-fluorouracil-treated MDSC: implication in cancer treatment. Cell Death & Disease, 10(7), 485.

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