Fig. 1.

Aflibercept forms 1:1 complexes with VEGF₁₆₅. The molar masses of aflibercept:VEGF₁₆₅ and bevacizumab:VEGF₁₆₅ complexes were analyzed by multi-angle laser light scattering detection coupled to SEC. The differential refractive index (right y axis) and the measured molar mass (left y axis) of peaks are indicated as a function of elution volume for each sample. The experimentally determined molar masses are indicated by horizontal lines. Cartoons of free VEGF₁₆₅ and complexes of aflibercept or bevacizumab bound to VEGF₁₆₅ are shown. Complexes of VEGF₁₆₅ with bevacizumab (a) or aflibercept (b) at various molar ratios were incubated for 12 h at ambient temperature. Following incubation, the samples were kept at 4 °C in the autosampler prior to injection (~100–200 µg per sample) onto a Superose 12 column pre-equilibrated in 10 mM phosphate containing 500 mM NaCl buffer (pH 7.0) with a flow rate of 0.3 mL/min. Chromatograms of VEGF₁₆₅ and bevacizumab (a) or aflibercept (b) are superimposed to indicate the elution profiles of the unbound proteins. The 1:1 molar ratio complexes yielded similar elution profiles and are not shown for the purposes of clarity