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Recombinant mouse il18

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Recombinant mouse IL18 is a cytokine that plays a role in the immune response. It is produced in a recombinant system and is suitable for use in research applications.

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Lab products found in correlation

Immunoprecipitation kit, by Promega (2 mentions) Immunoprecipitation kit, by Thermo Fisher Scientific (2 mentions) Il18 polyclonal antibody, by Abcam (2 mentions) Protein a agarose, by Thermo Fisher Scientific (2 mentions) Recombinant mouse il 12, by Thermo Fisher Scientific (2 mentions) Recombinant human il 15, by Miltenyi Biotec (1 mentions) Foxp3 transcription factor staining buffer set, by Thermo Fisher Scientific (1 mentions) Legend max elisa kit, by BioLegend (1 mentions)

4 protocols using recombinant mouse il18

1

Identification of IL18 Binding Proteins

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An immunoprecipitation kit (Promega, Madison, WI) was used to isolate IL18 binding proteins. Briefly, ECs from Il18r−/− mice were incubated with recombinant mouse IL18 (PeproTech, Inc., Rocky Hill, NJ, 50 ng/ml) for 15 minutes at 37 °C, washed with ice-cold PBS, lysed in a lysis buffer from the immunoprecipitation kit, and centrifuged at 14000 rpm for 15 minutes. The supernatant was passed through an IL18 polyclonal antibody (#5180R, BioVision, Inc., Milpitas, CA) pre-coated protein A agarose (Invitrogen) column, followed by extensive washes. Bound proteins were eluted according to the manufacturer’s instructions, and separated on an 8% SDS-PAGE. Silver staining displayed two bands of approximately 80-kDa and 125-kDa, which were removed and analyzed with mass spectrometry (Brigham and Women’s Hospital Core Facility).
Wang J., Sun C., Gerdes N., Liu C., Liao M., Liu J., Shi M.A., He A., Zhou Y., Sukhova G.K., Chen H., Cheng X., Kuzuya M., Murohara T., Zhang J., Cheng X., Jiang M., Shull G.E., Rogers S., Yang C.L., Ke Q., Jelen S., Bindels R., Ellison D.H., Jarolim P., Libby P, & Shi G.P. (2015). Interleukin 18 function requires both interleukin 18 receptor and Na-Cl co-transporter. Nature medicine, 21(7), 820-826.
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2

Measuring Intracellular IFNγ in Murine Splenocytes

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Spleens were processed and intracellular IFNγ measured as previously described.
49 (link) In short, splenocytes of 6–8‐week‐old mice were stimulated with 20 ng mL−1 recombinant mouse IL‐12 (Peprotech, Rehovot, Israel) in combination with 200 ng mL−1 recombinant mouse IL‐18 (Peprotech) or 50 ng mL−1 of recombinant human IL‐15 (Miltenyi Biotec, Bergisch Gladbach, Germany) for 4 h prior to fixation/permeabilization (Foxp3/Transcription Factor Staining Buffer Set; eBioscience™, San Diego, CA, USA) for detection of intracellular IFNγ by flow cytometry.
Meza Guzman L.G., Hyland C.D., Bidgood G.M., Leong E., Shen Z., Goh W., Rautela J., Vince J.E., Nicholson S.E, & Huntington N.D. (2023). CD45 limits early Natural Killer cell development. Immunology and Cell Biology, 102(1), 58-70.
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3

Identification of IL18 Binding Proteins

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An immunoprecipitation kit (Promega, Madison, WI) was used to isolate IL18 binding proteins. Briefly, ECs from Il18r−/− mice were incubated with recombinant mouse IL18 (PeproTech, Inc., Rocky Hill, NJ, 50 ng/ml) for 15 minutes at 37 °C, washed with ice-cold PBS, lysed in a lysis buffer from the immunoprecipitation kit, and centrifuged at 14000 rpm for 15 minutes. The supernatant was passed through an IL18 polyclonal antibody (#5180R, BioVision, Inc., Milpitas, CA) pre-coated protein A agarose (Invitrogen) column, followed by extensive washes. Bound proteins were eluted according to the manufacturer’s instructions, and separated on an 8% SDS-PAGE. Silver staining displayed two bands of approximately 80-kDa and 125-kDa, which were removed and analyzed with mass spectrometry (Brigham and Women’s Hospital Core Facility).
Wang J., Sun C., Gerdes N., Liu C., Liao M., Liu J., Shi M.A., He A., Zhou Y., Sukhova G.K., Chen H., Cheng X., Kuzuya M., Murohara T., Zhang J., Cheng X., Jiang M., Shull G.E., Rogers S., Yang C.L., Ke Q., Jelen S., Bindels R., Ellison D.H., Jarolim P., Libby P, & Shi G.P. (2015). Interleukin 18 function requires both interleukin 18 receptor and Na-Cl co-transporter. Nature medicine, 21(7), 820-826.
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4

Cytokine-Induced Secretion of IFN-γ and GM-CSF

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Around 2 × 10 4 mouse NK cells were stimulated with recombinant mouse IL-12 (20 ng ml -1 ; PeproTech, 210-12) and recombinant mouse IL-18 (10 ng ml -1 ; PeproTech) for 4 h before conditioned medium was harvested and stored at -80 °C. Mouse IFN-γ and GM-CSF were detected using Legend Max ELISA kits (BioLegend) following the manufacturer's instructions.
Cheng M.I., Li J.H., Riggan L., Chen B., Tafti R.Y., Chin S., Ma F., Pellegrini M., Hrncir H., Arnold A.P., O'Sullivan T.E, & Su M.A. (2023). The X-linked epigenetic regulator UTX controls NK cell-intrinsic sex differences. Nature immunology, 24(5).
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