Fig. 4

In vitro characterization of anti-O antigen mAbs. a Association/dissociation curves of K. pneumoniae O1 or K. pneumoniae O2 LPS binding to mAbs KPE33, KPN70, and KPN42 were measured by ForteBio Octet using protein A capture probe pre-loaded with each mAb. b Confocal images of mAb binding to an O1 (Kp43816) or O2 (Kp9148) strain (scale bar, 2 μm). c LPS neutralization was measured using RAW264.7 cells transfected with a NK-kB driven luciferase promoter stimulated with 2 ng ml⁻¹ of purified O1 or O2 LPS in the presence of KPE33, KPN70, KPN42 or the isotype-matched control mAb (c-IgG). Percent inhibition was calculated as the amount of luminescence (RLU) in treated wells divided by the RLU in wells stimulated with LPS alone multiplied by 100. d Opsonophagocytic killing assays were performed using various concentrations of mAbs in the presence of the phagocytic cell line HL-60. The capsule-deficient luminescent mutants Kp43816ΔcpsB and Kp8570ΔcpsB were used as the O1 and O2 serotype targets, respectively. Percent killing was calculated based on the amount of luminescence after 2.5 h incubation in treated wells vs. wells with bacteria only. An isotype-matched mAb (c-IgG) is used as a negative control. e Summary of the in vitro functional activities against O1 and O2 K. pneumoniae for each mAb. All error bars indicate s.d. at each data point