Assessing the stability of pre-F during FI-RSV production.
(a) Dot blots demonstrate motavizumab recognition of antigenic site II, AM14 recognition of quaternary antigenic site V and D25 and 5C4 recognition of antigenic site Ø after RSV is incubated at 37 °C with a 0.025% concentration of formalin in 24-hour intervals. Average antibody binding of virions is reported relative to antibody binding of a pre-F protein loading control with error bars representing standard deviation of three experiments. Significant differences between populations are calculated by One-Way ANOVA and noted graphically (ns, not significant; *p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001; ****p ≤ 0.0001). Recognition of antigenic site II, present on both pre-F and post-F surfaces, is preserved after treatment with high temperature and a dilute concentration of formalin. In contrast, dot blots demonstrate pre-F-specific mAbs AM14, D25 and 5C4 rapidly lose reactivity against virions incubated at 37 °C with a dilute concentration of formalin. (b) Percentage of cells infected by RSV incubated at 37 °C with a 0.025% concentration formalin in 24-hour intervals by flow cytometry (Chen 2010).