Robustness of iCIEF methodology for the analysis of monoclonal antibodies: An interlaboratory study

Abstract

An international team including 12 laboratories from 11 independent biopharmaceutical companies in the United States and Switzerland was formed to evaluate the precision and robustness of imaged capillary isoelectric focusing for the charge heterogeneity analysis of monoclonal antibodies. The different laboratories determined the apparent pI and the relative distribution of the charged isoforms for a representative monoclonal antibody sample using the same capillary isoelectric focusing assay. Statistical evaluation of the data was performed to determine within and between laboratory consistencies and outlying information. The apparent pI data generated for each charged variant peak showed very good precision between laboratories with RSD values of less than 0.8%. Similarly, the RSD for the therapeutic monoclonal antibody charged variants percent peak area values are less than 11% across different laboratories using different analyst, different lots of ampholytes and multiple instruments. These results validate the appropriate use of imaged capillary isoelectric focusing in the biopharmaceutical industry in support of process development and regulatory submissions of therapeutic antibodies.

Evaluation of capillary zone electrophoresis for charge heterogeneity testing of monoclonal antibodies

“…As a result approx. 20% of the whole dataset was removed, which is a similar proportion of outliers found and removed throughout the evaluation of other methods for determination of charge heterogeneity [23].…”

“…An international team of 11 laboratories from 9 independent companies/organizations in the US, Switzerland and Germany was formed to contribute to this study. As already demonstrated for capillary IEF [22] and imaged capillary IEF [23] the aim of the present CZE study was to show applicability in this context.…”

Recent advances in the analysis of therapeutic proteins by capillary and microchip electrophoresis

“…The innovator, the biosimilar manufacturer, and the regulatory agency need to be certain that, despite the various laboratory conditions, the experimental results are comparable. To help facilitate this, Salas-Solano et al evaluated an iCIEF method in 12 different laboratories across the world using several analysts, a variety of ampholytes, and multiple instruments [128]. The combined precision for the 12 labs was 0.8% RSD for the pI determination and 11% RSD for the percent peak area values for the charge variants of a therapeutic mAb.…”

The Use of Electrophoresis for the Study of Saliva Involvement in Ingestive Behavior

“…Capillary isoelectric focusing is a type of CE that offers significant advantages over isoelectric electrophoresis slab gels in terms of automation, separation speed and quantitation. Although at the beginning capillary isoelectric focusing was not readily accepted, due to the difficulties in isolating the focused zones for the detector, actually this separation technique presents the highest resolution of all charge based separation techniques, due to nowadays systems that allow the monitorization of the whole column at once [90]. Micellar electrokinetic capillary chromatography is an electrophoretic technique developed in the early 90's that extends the applicability to non-charged molecules, which cannot be separated using simple free solution CE [91].…”