Development and Optimization of a LabChip® Method for Characterization of a Heavily Sialylated Protein and Its Advantage in High-Throughput InProcess Development Support

Pharmaceutical process development generates thousands of samples in crude matrix within a very short
period of time, which requires rapid result turnout from a robust high-throughput analytical method.
Microchip-based gel electrophoresis was evaluated as a high-throughput alternative to conventional capillary
electrophoresis sodium dodecyl sulfate (CE-SDS) for monitoring quality attributes of process development
samples. Separation in CE-SDS is based on apparent molecular size. One application is the separation and
quantitation of the degree of glycosylation under reducing conditions.
In this study, HT LabChip® GXII Touch system was used to monitor the aglycosyated species for a heavily
sialylated protein. Some challenges were encountered using the standard conditions with Protein Express
LabChip® for this heavily sialylated protein. A progressively deteriorated electrophoretic profile was
observed with the accumulation of replicate injections, for the same sample preparation (< n=10). While
investigating the atypical profiles, several critical parameters were identified and optimized to obtain stable
and reproducible resolution. In this presentation, we will discuss those critical parameters which are
important for separation of heavily sialylated proteins using the LabChip® system.
Additionally, the optimized method was further evaluated for specificity, linearity, precision, and limit of
quantitation, as compared with conventional CE-SDS. The LabChip® method has been established for
monitoring aglycosylated species for this heavily sialylated protein, in either pure or crude in-process
sample media with sufficient resolution and sensitivity, but on a time scale of approximately 20 times faster
(2 min versus 40 min per sample) than conventional CE-SDS separations.