Figure 1.
SAR443809 binds to FBb but not FB. (A) Target binding kinetics of SAR443809. The binding affinity of SAR443809 to human FB (orange) or human FBb (red) was assessed using surface plasmon resonance (Biacore T200). The mAb was captured on a Series S Sensor Chip Protein A and the kinetic binding constants were obtained in single-cycle kinetics mode with increasing concentrations of the targets. Data were analyzed using the Biacore T200 Evaluation Software, and ka, kd, and KD values were calculated based on a 1:1 binding model curve fit to double reference-subtracted sensorgrams (black). (B) SAR443809 Fab binding to AP C3 convertase (C3bBb). The binding affinity of SAR443809 Fab to C3bBb (red) was assessed using surface plasmon resonance (Biacore T200). A small amount of C3b was amine coupled on a Series S CM5 chip, followed by natural C3b cleavage and subsequent covalent attachment to the chip via C3bBb formed on the surface (multiple injections of FB, FD, and C3). After surface preparation, C3bBb was formed via injection of FB and FD, and kinetic data were collected in single-cycle kinetics mode with increasing concentration of SAR443809 Fab. Data were analyzed using the Biacore T200 Evaluation Software, and ka, kd, and KD values were calculated based on a 1:1 binding model curve fit to double reference-subtracted sensorgrams (black). (C) SAR443809 interaction with FB in solution. The mass distribution of samples containing either SAR443809 (orange), FB (blue), or a 2:1 FB:SAR443809 mixture (green) was measured using mass photometry. Histograms for individual components reveal expected molecular weights (SAR443809-146 kDa; FB – 93 kDa). The mixture of SAR443809 and FB shows 2 peaks corresponding to the mass of the individual species (SAR443809-146 kDa; FB – 93 kDa), with no higher molecular weight species present. (D) SAR443809 interaction with FBb in solution. The mass distribution of samples containing either SAR443809 (orange), FBb (blue), or a 2:1 FBb:SAR443809 mixture (green) was measured using mass photometry. Histograms for individual components reveal expected molecular weights (SAR443809-146 kDa; FBb – 60 kDa). The mixture of SAR443809 and FBb shows 4 peaks, corresponding to free FBb (60 kDa), free SAR443809 (146 kDa), SAR443809-FBb1 complex (206 kDa), and SAR443809-FBb2 complex (266 kDa).

SAR443809 binds to FBb but not FB. (A) Target binding kinetics of SAR443809. The binding affinity of SAR443809 to human FB (orange) or human FBb (red) was assessed using surface plasmon resonance (Biacore T200). The mAb was captured on a Series S Sensor Chip Protein A and the kinetic binding constants were obtained in single-cycle kinetics mode with increasing concentrations of the targets. Data were analyzed using the Biacore T200 Evaluation Software, and ka, kd, and KD values were calculated based on a 1:1 binding model curve fit to double reference-subtracted sensorgrams (black). (B) SAR443809 Fab binding to AP C3 convertase (C3bBb). The binding affinity of SAR443809 Fab to C3bBb (red) was assessed using surface plasmon resonance (Biacore T200). A small amount of C3b was amine coupled on a Series S CM5 chip, followed by natural C3b cleavage and subsequent covalent attachment to the chip via C3bBb formed on the surface (multiple injections of FB, FD, and C3). After surface preparation, C3bBb was formed via injection of FB and FD, and kinetic data were collected in single-cycle kinetics mode with increasing concentration of SAR443809 Fab. Data were analyzed using the Biacore T200 Evaluation Software, and ka, kd, and KD values were calculated based on a 1:1 binding model curve fit to double reference-subtracted sensorgrams (black). (C) SAR443809 interaction with FB in solution. The mass distribution of samples containing either SAR443809 (orange), FB (blue), or a 2:1 FB:SAR443809 mixture (green) was measured using mass photometry. Histograms for individual components reveal expected molecular weights (SAR443809-146 kDa; FB – 93 kDa). The mixture of SAR443809 and FB shows 2 peaks corresponding to the mass of the individual species (SAR443809-146 kDa; FB – 93 kDa), with no higher molecular weight species present. (D) SAR443809 interaction with FBb in solution. The mass distribution of samples containing either SAR443809 (orange), FBb (blue), or a 2:1 FBb:SAR443809 mixture (green) was measured using mass photometry. Histograms for individual components reveal expected molecular weights (SAR443809-146 kDa; FBb – 60 kDa). The mixture of SAR443809 and FBb shows 4 peaks, corresponding to free FBb (60 kDa), free SAR443809 (146 kDa), SAR443809-FBb1 complex (206 kDa), and SAR443809-FBb2 complex (266 kDa).

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