Abstract
A capillary zone electrophoresis (CZE) method was developed for the rapid analysis of charge heterogeneity of immunoglobulin G (IgG) monoclonal antibodies (mAbs). The separation was carried out in a short, dynamically coated fusedโsilica capillary. A number of separation parameters were investigated and optimized, including pH, concentration of the separation buffer (ฮตโamino caproic acid), concentration of the triethylenetetramine (TETA) dynamic coating, the capillary internal diameter and the field strength used for the separation. The effects of betweenโrun flushing of the capillary and the data acquisition rate were also evaluated. Under the optimized conditions, a fast (<5โmin), selective and reproducible separation of mAb charge variants was achieved under a very high electric field strength (1000โV/cm). This method also requires only a short conditioning of the capillary, with betweenโrun conditioning completed within 2โmin. The method was evaluated for specificity, sensitivity, linearity, accuracy and precision. The same separation conditions were applied to the rapid separation (2โ5โmin) of charge variants of multiple monoclonal antibodies with p__I__ in the range of 7.0โ9.5. Compared with other existing methods for charge variants analysis, this method has several advantages including a short run time, rapid capillary conditioning and simple sample preparation.