Macromolecules such as therapeutic proteins currently serve an important role in the treatment of eye diseases such as wet age-related macular degeneration and diabetic retinopathy. Particularly, bevacizumab and ranibizumab have been shown to be effective in the treatment of these diseases. Iontophoresis can be employed to enhance ocular delivery of these macromolecules, but the lack of information on the properties of these macromolecules has hindered its development. The objectives of the present study were to determine the effective electrophoretic mobilities and charges of bevacizumab, ranibizumab, and model compound polystyrene sulfonate (PSS) using capillary zone electrophoresis. Salicylate, lidocaine, and bovine serum albumin (BSA), which have known electrophoretic mobilities in the literature, were also studied to validate the present technique. The hydrodynamic radii and diffusion coefficients of BSA, bevacizumab, ranibizumab, and PSS were measured by dynamic light scattering. The effective charges were calculated using the Einstein relation between diffusion coefficient and electrophoretic mobility and the Henry equation. The results show that bevacizumab and ranibizumab have low electrophoretic mobilities and are net negatively charged in phosphate buffered saline (PBS) of pH 7.4 and 0.16M ionic strength. PSS has high negative charge but the electrophoretic mobility in PBS is lower than that expected from the polymer structure. The present study demonstrated that capillary electrophoresis could be used to characterize the mobility and charge properties of drug candidates in the development of iontophoretic drug delivery.