Il 10 elisa kit

Serum samples of immunized mice and naïve mice were gathered for cytokines and antibody levels detection 30 or 75‐days post‐infection. The effect of vaccine‐induced cytokine production was measured using mouse IFN‐γ, TNF‐α, IL‐12p70 and IL‐10 ELISA kits (BioLegend, San Diego, CA, USA) according to the manufacturer’s protocols. The levels of T. gondii specific total immunoglobulin G (IgG) and subclasses of IgG antibodies IgG2a and IgG1 as indicators of type 1T‐helper (Th1) and type 2T‐helper (Th2) responses, respectively, were analysed using ELISA assay as described previously (Wang et al., 2017a; Huang et al., 2019). A total of 100 μl soluble tachyzoite antigens (5 μg ml⁻¹) were coated in a 96‐well plate and incubated at 4°C overnight. Then, the plate was washed 3 times and blocked with 1% BSA at 37°C for 1 h. After washing 3 times, 100 μl serum samples were added to appropriate wells at 37°C for 45 min. Subsequently, 100 μl of HRP conjugated goat anti‐mouse IgG secondary antibodies (1:5000 dilutions, Beyotime Biotechnology, Beijing, China), anti‐mouse IgG1(1:10 000, Abcam, Cambridge, UK) and anti‐mouse IgG2a (1:10 000, Abcam) were added to each well and incubated at 37°C for 30 min. Finally, tetramethylbenzidine as substrate was incubated to visualize the immunoreaction, and the OD value was determined at 630 nm using a microplate reader.

IDCs were co-cultured with killed tumor cells (as indicated above) or stimulated with lipopolysaccharide (LPS Cat# L2630) as a positive control. The culture supernatant was then harvested to detect the level of IL-12p70 and IL-10 secretion using ELISA kit (IL-12p70 ELISA kit (BioLegend Cat#431704); IL-10 ELISA kit (BioLegend Cat# 430604)) according to the manufacturer’s instructions. After co-culture with cancer cells unpulsed or tumor-loaded DCs were added to autologous T cells at a ratio of 1:10. After 48 hours, the production of IFN-γ was measured by ELISA (BioLegend Cat# 430104) according to the manufacturer’s instructions.