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2 protocols using recombinant tnf α il 6 il 8

1

Immune Cell Signaling Assay Reagents

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Compound 48/80, 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT), phorbol 12-myristate 13-acetate (PMA), and calcium ionophore A23187 (A23187) were purchased from Sigma-Aldrich Chemical Co. (St. Louis, MO, USA). 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt (MTS) was purchased from Promega Corporation (Madison, WI, USA). Avidin peroxidase (AP), fetal bovine serum (FBS), RPMI 1640, and Iscove's Modified Dulbecco's Medium (IMDM) were purchased from Gibco BRL (Grand Island, NY, USA). Anti-human TNF-α/IL-6/IL-8, anti-mouse immunoglobulin E (IgE)/IL-6, recombinant TNF-α/IL-6/IL-8, and biotinylated TNF-α/IL-6/IL-8 were purchased from BD Pharmingen (San Diego, CA, USA). Anti-phospho-p38, -ERK, -JNK, -Akt, -IκBα, anti-GAPDH, IκBα, and NF-κB antibodies were purchased from Cell Signaling Technology, Inc. (Danvers, MA, USA). Anti-p38, JNK, and histone antibodies were purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA, USA). Anti-Akt and ERK antibodies were purchased from Bioworld Technology (Minneapolis, MN, USA).
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2

Inflammatory Signaling Pathway Analysis

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2, 4-dinitrofluorobenzene (DNFB), compound 48/80, 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT), PMA, and A23187 were purchased from Sigma-Aldrich Chemical Co. (St. Louis, MO, USA). Avidin peroxidase (AP), fetal bovine serum (FBS), Roswell Park Memorial Institute (RPMI) 1640, and Iscove's Modified Dulbecco's Medium (IMDM) were purchased from Gibco BRL (Grand Island, NY, USA). Anti-human TNF-α/IL-6/IL-8, recombinant TNF-α/IL-6/IL-8, biotinylated TNF-α/IL-6/IL-8, anti-mouse IgE/IL-1β, recombinant IgE, IL-1β, biotinylated IgE, and IL-1β were purchased from BD Pharmingen (San Diego, CA, USA). Anti-phospho p38 antibody was purchased from Cell Signaling Technology, Inc. (Danvers, MA, USA). Anti-phospho ERK, JNK, IκBα, anti-p38, JNK, NF-κB, histone, and GAPDH antibodies were purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA, USA). Anti-ERK, and α-tubulin antibodies were purchased from Bioworld Technology (Minneapolis, MN, USA).
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