The single primary hepatocytes were pipetted under fluorescence microscope and seeded in one well of 96-well plates. After cultured in IL6-HCM for 14 days, the single primary hepatocytes derived IL6-iHPC clones were trypsinized and reseeded into new 12-well plate for further expanding. Quantita...
Neutrophils were incubated in duplicate wells of flat bottom 96-well plates containing hyphae that had been grown for 3 h with or without serum opsonization (2% heat-inactivated mouse serum; Gemini Bio-Products), at a neutrophil to C. albicans ratio of 1:4 at 37 °C. After 2.5 ...
Based on previous reports (Kimura et al., 2014; Saito et al., 2011), 2H4 cells (2 × 105 cells/50 μl/well) in 96-well black plates (Greiner Bio-One GmbH, Frickenhausen, Germany) were pretreated with different concentrations of individual chemicals for 1 h. The 2H4 cells were then...
For the reporter gene assays, 293 cells (1.6 × 105 cells/well) were seeded into six-well (35 mm) plates. At 24 hr after seeding, cells were transfected with a reporter gene plasmid and each expression plasmid as indicated. An Ig-κ-luciferase reporter was used to measure NF-κB-depende...
Briefly, after coating microtitre plates (NUNC, Thermo Scientific, Waltham, MA, USA) with purified monoclonal antibodies reactive to mouse cytokines IL-10, TNF-α, IL-6, IL-17 (BD, New Jersey, USA), standards and samples were added and incubated overnight at 4°C. Biotinylated monoclonal ...
Interferons (IFNs) are critical soluble factors in the immune system and are composed of three types, (I, II and III) that utilize different receptor complexes IFN-αR1/IFN-αR2, IFN-γR1/IFN-γR2, and IFN-λR1/IL-10R2, respectively. Here we identify IFN-υ from the genomic sequence...
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Nunc MaxiSorp and PolySorp ELISA plates (Nunc A/S, Roskilde, Sjælland, Denmark) were coated with IL-22R1 (2.5 µg/ml). Then, MaxiSorp plates were blocked with Protein-Free (PBS) Blocking Buffer and PolySorp plates were blocked with 1% BSA in PBS-T (PBS, 0.05% Tween-20). ...
Severe infection commonly results in immunosuppression, which leads to impaired pathogen clearance or increased secondary infection in both humans and animals. However, the exact mechanisms remain poorly understood. Here, we demonstrate that IL-33 result