Fig. 2: In vitro binding, kinetics and neutralization data for PMD based peptides.

a Scatter plot depicting peptide-mediated binding potencies determined using an AlphaLISA competition binding assay (ALC), reported as log IC₅₀ in molar (x-axis) versus neutralization activity determined using virus neutralization assay (VNA), reported as log EC₅₀ in molar (y-axis). ALC and VNA assays were performed using H1N1 A/California/07/2009 (H1/Cal), H1N1 A/New Caledonia/20/1999 (H1/NCa), and H5N1 A/Vietnam/1203/2004 (H5/Viet) HAs and corresponding H1N1 and H5N1 virus strains respectively. Competition against the HA stem-binding protein HB80.4³⁴ and Fab of HA head binding antibody 2D1⁸⁰ were performed in the ALC assay. Each dot represents one tested peptide. The most potent peptides are highlighted as CP1, yellow; CP4, magenta, and CP8, green, whereas the non-template-based peptide (CP14, red) and other weak binding peptides are represented as black dots. Peptide CP5 was not included in the analysis. The dotted line represents the lower limit of quantification in the assays. b Dose-dependent competitive binding curves for peptide CP1 binding to the H1/Cal, H1/NCa and H5/Viet HAs, respectively. HA stem binding small protein HB80.4 and Fab of HA head binding antibody 2D1 used in the ALC assay. Peptide concentrations are reported in log molar (x-axis) versus % normalized binding response (y-axis). c Surface plasmon resonance (SPR) derived kinetic data for the peptides, as determined for H1/Cal, H1/NCa and H5/Viet HAs. Peptide half-life is calculated as t_1/2 = ln 2/ k_off. Values reported in the table are an average of two independent experiments. Errors are calculated as uncertainty in the mean, \(\Delta {{\mathrm{x}}}_{{avg}}=R / (2\sqrt{N})\); where, x = experimental data values, R= difference between the maximum and minimum values of x. N = number of measurements. SPR measurements were not performed for peptide CP12. SPR response is measured in response units (RU). d Statistical comparison of binding data of peptides in surface plasmon resonance (SPR) and AlphaLISA assay.