Development of imaged capillary isoelectric focusing method and use of capillary zone electrophoresis in hepatitis B vaccine RECOMBIVAX HB®.

Abstract

The hepatitis B virus vaccine consists of a major surface antigen called HBsAg, which is a lipid-bound protein that self-assembles into 22 nm spherical noninfectious virus-like particles (VLPs). The HBsAg VLP particles are expressed in yeast and have been well-characterized biochemically and biophysically employing various analytical techniques. In fact, a CZE method has been developed for monitoring process purification of the hepatitis B vaccine. Another CE-based method, imaged capillary IEF (icIEF) has been used extensively in the field of protein-based drug development as a tool for product identification,stability monitoring, and characterization. Here we describe the development of the icIEF method using the iCE280 instrument from ProteinSimple for measuring the pI and monitoring the profiles of HBsAg VLP particles. This method was applied to characterize the stability of the HBsAg VLP particles in three different formulation buffers. The results show that HBsAg VLP particles have a pI of 2.7 and it is one of most acidic particles that we have measured by icIEF. In addition to icIEF, we have also employed a CZE method to measure the electrophoretic mobility of HBsAg VLP particles and compared the results with icIEF and dynamic light scattering methods, showing consistent correlation among the three methods in terms of HBsAg VLP particles aggregation.