Journal: NPJ Vaccines

Article Title: Adenovectors encoding RSV-F protein induce durable and mucosal immunity in macaques after two intramuscular administrations

doi: 10.1038/s41541-019-0150-4

Figure Lengend Snippet: RSV-F-specific CD8+ T cells a – d and CD4+ T cells e–h expressing IFNγ a , e , TNFα b , f , or IL-2 c , g were determined in cells extracted from BAL, using polychromatic flow cytometry, after stimulation with a pool of 15-mer peptides overlapping by 11 amino acids, covering the RSV-F protein sequence. Shown are arithmetic group means with upper 95% confidence limit (upper whisker) over time, of individually background subtracted values. Animals received a homologous prime-boost with the Ad26/Ad35 mix (light gray), a homologous prime-boost with Ad26 (dark gray), or a heterologous prime-boost with Ad26 and Ad35 (black). Arrows indicate immunization time points. Polyfunctional composition of the cytokine response in CD8+ T cells e and CD4+ T cells h of individual animals 4 weeks post boost (study week 16) was determined as described before and is summarized in the pie charts. Responders as defined by Fisher’s exact test are indicated by an asterisk. Data from individual animals in Figures d , h are shown in corresponding order.

Article Snippet: Cells were washed in FACS staining buffer, then surface stained for 30 min at RT in the dark with 100 µl of a cocktail containing antibodies targeting CD3 (BD Biosciences, clone SP34-2, catalog number 557917, 1 µl), CD4 (BD Biosciences, catalog number 562843, clone L200, 0.5 µl), CD8 (BD Biosciences, catalog number 557834, clone SK1, 5 µl), CD28 (BD Biosciences, clone L293, catalog number 337181, 20 µl), and CD95 (clone DX2, catalog number 561978, 20 µl).

Techniques: Expressing, Flow Cytometry, Sequencing, Whisker Assay