Apc conjugated anti cd11c antibody

Fitc-conjugated anti-CD4 antibody, PE-conjugated anti-IFNγ antibody, APC-conjugated anti-IL4 antibody, PE/Cy7-conjugated anti-IL17 antibody, and Percp/Cy5.5-conjugated anti-CD8 antibody were purchased from ebioscience. Biotin-conjugated PNA was from Vector lab. APC-conjugated anti-GL7 antibody, PE-conjugated anti-PD-1antibody, APC-conjugated anti-CXCR5 antibody, PE-conjugated anti-CD169 antibody, APC-conjugated anti-CD11c antibody, PE/Cy7-conjugated anti-CD14 antibody, Percp/cy5.5-conjugated anti-CD19 antibody, pacific blue-conjugated anti-IgD antibody, PE-conjugated anti-CD21/35 antibody, PE-conjugated anti-CD19 antibody, Alex Fluor@488-conjugated streptavidin, PE-conjugated streptavidin, Fitc-conjugated anti-CD80 antibody, and PE/Cy7-conjugated anti-CD86 antibody were obtained from biolegend. Mouse IL4, IL17, IL10, and IFNγ ELISA kit were bought from Neobioscience. Lymphocytes Separation Medium was purchased from MP Biomedicals, LLC. Micro-osmotic pump for 14 days was from DURECT Corporation (Cupertino, CA, USA). MultiScreen@HTS96-well plates were obtained from EMD Millipore Corporation (Billeria, MA, USA).

Phenotypic maturation of DCs was analyzed by flow cytometry, cells were resuspended in PBS containing 2% FBS and stained with APC-conjugated anti-CD11c antibody, PC7-conjugated anti-CD40, PC5.5-conjugated anti-MHCII, FITC-conjugated anti-CD83 and PE-conjugated anti-CD86 anti-body (Biolegend, CA, USA). Non-treated DCs were also stained with isotype IgG as negative control. To collect DCs and CD3 co-cultured cells, splenocytes from the same mouse strain were obtained, and incubated with anti-CD3 beads followed by magnetic sorting (MACS). DC at 2 × 10⁴ cells/well from the BM of C57BL/6J were seeded in 96-well round-bottom plates, pulsed with Lewis cell lysates (5 μg/ml) and incubated with YYWY (50 or 100 μg/ml), LPS or un-treated at 5μg/ml was added and incubated with cells for 24 h at 37°C. CD3 cells per well and co-cultured with DC (2 × 10⁴/well) in 96-well round-bottom plate for 96 h. Then, the surface markers were stained with FITC-conjugated anti-CD8 (Biolegend, CA, USA), before lysis of the cell membrane with BD Cytofix/Cytoperm™ ContentsFixation, then the surface markers were stained with FITC-conjugated anti-CD8 (Biolegend, CA, USA), before lysis of the cell membrane with BD.