Anti cd45ra pacific blue

Four days after infection, T cells were harvested and washed once with PBS, stained with rabbit anti-FLT3L antibody (Abcam, USA) for 1 h at 4 °C, and washed twice. Then PE donkey anti-rabbit IgG antibody (Biolegend, USA) was added, incubated at 4 °C for 30 min, and analyzed by flow cytometry using CantoII flow cytometer (BD Biosciences, San Jose, CA, USA) [14 (link)]. For T cell phenotype analysis, T cells were harvested 7 days after infection and washed once with PBS, stained with anti-CD4-PE/Cy7 (Biolegend, USA), anti-CD8-PerCP-Cy5.5 (Biolegend, USA), anti-CCR7-PE (Biolegend, USA), and anti-CD45RA-Pacific Blue (Biolegend USA) 30 min at 4 °C, then washed and resuspended in PBS for flow cytometry analysis [15 (link)].

Phenotypic characteristics of immune cells were assessed by flow cytometry (FACSVerse cytometer; BD, USA) using monoclonal antibodies: anti-CD25-FITC, anti-CD45R0-FITC, anti-CD19-PE-Cy7, anti-CD4-PE-Cy7, anti-CD3-PE-Cy7, anti-CD8-PE-Cy7, anti-CD127-PE-Cy7, anti-CD14-PerCP, and anti-CD45RA-Pacific Blue (Biolegend, USA); and anti-TNFR1-PE, anti-TNFR2-PE, anti-TNFR1-APC, and anti-TNFR2-APC (R&D Systems, USA). Data processing and calculation of fluorescence intensity parameters were performed in the FacsDiva software (BD, USA). To calculate the number of receptor molecules per cell, the BD QuantiBRITE PE Kit (BD Biosciences, USA) was used according to the method described earlier [22 (link)].