Mouse il 10 flex set

Purified B cells were cultured with LPS (10 µg/ml) for 16 h. Concentration of secreted cytokines was measured from supernatants using mouse IL-10 Flex Set (BD) and mouse IL-6, TNFa FlowCytomix Simplex (Bender MedSystem).

The contents of IL-10 (Mouse IL-10 Flex Set, 558300, BD), IFN-γ (Mouse IFN-γ Flex Set, 558296, BD), TNF (Mouse TNF Flex Set, 558299, BD), and IL-2 (Mouse IL-2 Flex Set, 558297, BD) in the serum of mice were detected using CBA according to the manufacturer’s protocols. Initially, the different concentrations of standard products were formulated according to the respective preparations, and a standard curve was plotted. Then, 50 μL of serum and 50 μL of microspheres were mixed and incubated at room temperature in the dark for 2 h. Then, 50 μL of PE antibody conjugated with TNF, IL-2, IL-1β, and IFN-γ was added and incubated for 1 h at room temperature in the dark. Further, 1 mL of washing solution was added and centrifuged, and the supernatant was gently discarded. The resulting suspension was resuspended in 400 μL of the buffer, and finally, the cytokine levels were measured by FCM, and the results were analyzed using the Diva (BD).