Rat th1 th2 cytokine assay kit

Cytokine expression in the peripheral blood and synovial fluid was detected by flow cytometry using a rat Th1/Th2 Cytokine Assay kit (BioLegend). The magnetic beads, assay buffer, antibody, and standard or tested sample were incubated together for 3 h at room temperature. The concentrations of granulocyte-macrophage colony-stimulating factor (GM-CSF), IFN-γ, interleukin (IL)-2, IL-4, IL-5, IL-10, IL-6, IL-13, and TNF-α in rat serum and synovial fluid were measured with flow cytometry.

Peripheral blood samples were collected and centrifuged at 10,000 rpm for 20 min. The serum levels of interleukin-2 (IL-2), interleukin-4 (IL-4), interleukin-5 (IL-5), interleukin-6 (IL-6), interleukin-10 (IL-10), interleukin-13 (IL-13), granulocyte-macrophage colony-stimulating factor (GM-CSF), interferon-gamma (IFN-γ) and tumor necrosis factor alpha (TNF-a) were measured using a rat Th1/Th2 cytokine assay kit (BioLegend). Each well of the detection plate was supplied with the capture beads, antibody, fluorescent reagent and the standard or tested sample, and the plate was incubated for 3 h at 4°C on a shaker at a speed of 500 rpm in the dark. Cytokines were analyzed using NovoCyte flow cytometry (American ACEA BIO, NovoCyte D2040R). Data were analyzed using LEGENDplex v8.0 (BioLegend).
Th1 (IFN-γ and IL-2) and Th2 (IL-4, IL-5 and IL-13) cytokines were measured using flow cytometry analysis. The Th1/Th2 ratio was calculated based on the average fluorescence intensities of these cytokines and referred to the study by Anand (34 ).