Fig 2: Platelets bind and internalize RSV particles, which does not lead to platelet degranulation in vitro. (a) Washed platelets were incubated for 1 h with sucrose-purified RSV-A2 at the indicated concentrations and analysed by flow cytometry after staining with an FITC-conjugated antibody specific for the RSV attachment protein G (131-2G). Graphs show the percentage of FITC-positive platelets of different donors (n=4–6). (b) Washed platelets were incubated for 1 h with 0.1 PFUVero per platelet of RSV strains A2 (ATCC VR-1540) or X (GenBank FJ944820.1) and analysed as in (a). (c) Following 1 h of incubation with RSV-A2 at 0.1 PFUVero per platelet, platelets were fixed and permeabilized with a cytofix/cytoperm kit, stained as in (a) and measured with flow cytometry to assess RSV internalization. Following 1 h of incubation with RSV-A2 at 0.1 PFUVero per platelet, platelet degranulation was assessed by measuring P-selectin (d) and CD63 (e) surface expression and CCL5 secretion (f). Each data point indicates the average of technical duplicates from a single platelet donor. Abbreviations: perm, permeabilized; PFU, plaque-forming units; suc, sucrose control; TRAP-6, thrombin receptor activating peptide-6.