Intercompany Study to Evaluate the Robustness of Capillary Isoelectric Focusing Technology for the Analysis of Monoclonal Antibodies

Interlaboratory comparisons are essential to bringing emerging technologies into biopharmaceutical industry practice and regulatory acceptance. As a result, an international team including 12 laboratories from 10 independent biopharmaceutical companies in the United States and Switzerland was formed to evaluate the precision and robustness of capillary isoelectric focusing (CIEF) to assess the charge heterogeneity of monoclonal antibodies. The different laboratories determined the apparent pI and the relative distribution of the charge isoforms of a representative monoclonal antibody (rMAb) sample using the same CIEF method. Statistical evaluation of the data was performed to determine within and between-laboratory consistencies and outlying information. The apparent pI data generated for each charge variant peak showed very good precision between laboratories with percentage of RSD values of ??0.5%. Similarly, the percentage of RSD for the rMAb charge variants percent peak area values are ??4.4% across different laboratories with different analysts using different lots of ampholytes and multiple instruments. Taken together, these results validate the appropriate use of CIEF in the biopharmaceutical industry in support of regulatory submissions.     

A Series of Collaborations Between Various Pharmaceutical Companies and Regulatory Authorities Concerning the Analysis of Biomolecules Using Capillary Electrophoresis

An international project team (including members from US, Canada and UK) has been formed from a number of interested biopharmaceutical companies and regulatory authorities to conduct a cross-organisation collaboration exercise. The results from this exercise demonstrate the robustness of CE-SDS across eight different organisations that used instruments of the same equipment model, the same reagents, and the same methodology. Data generated from the analysis of a series of molecular weight markers showed very good precision with regards to relative migration time (RMT) both within and between organisations. The apparent molecular weight of bovine serum albumin (BSA) was measured with good precision to within approximately 2% RSD across the participants. A representative IgG sample showed similar results with regards to relative migration time of its 3 main components, IgG Light Chain, IgG Non-glycosylated Heavy Chain, and IgG Heavy Chain. Fractional peak area for each peak also showed good agreement, with less than 9% RSD for all peaks. This exercise will facilitate both increased regulatory and industrial opinion of CE for biopharmaceutical analysis.CE in the Biotechnology & Pharmaceutical Industries: 7th Symposium on the Practical Applications for the Analysis of Proteins, Nucleotides and Small Molecules, Montreal, Canada, August 12–16, 2005     

A Series of Collaborations between Various Pharmaceutical Companies and Regulatory Authorities Concerning the Analysis of Biomolecules Using Capillary Electrophoresis: Additional Instruments/Buffer

An international project team (including members from US, Canada and UK) was formed from a number of interested biopharmaceutical companies and regulatory authorities to conduct a cross-organisation collaboration exercise. The results of the first comparison with eight different organisations that used instruments of the same equipment model, the same reagents, and the same methodology has been reported previously [1]. This report represents the addition of other instruments using a different run buffer. The relative migration times were different, as expected, prohibiting a direct comparison between companies. The within-organisation variability was low for both relative migration time (<0.34% RSD% for all companies save one) and the peak area (<5% RSD% for all companies save one) when measuring the purity of a representative IgG sample. The apparent molecular weight of bovine serum albumin was measured with good precision (less than 10% RSD% across all companies) to the theoretical value when all data is utilized (67.5 kDa compared to 66.4 kDa). For a representative IgG sample, the three main components, IgG Light Chain, IgG Non-glycosylated Heavy Chain, and IgG Heavy Chain, could not be separated, specifically the IgG Non-glycosylated Heavy Chain and IgG Heavy Chain. When the IgG Non-glycosylated Heavy Chain and IgG Heavy Chain were combined for all organisations, the fractional peak area for the IgG Light Chain and IgG Non-glycosylated Heavy Chain + IgG Heavy Chain peak also showed excellent agreement, with less than 7.5 and 3.5% RSD%, respectively. The value of this exercise is in demonstrating the reliability of CE for the determination of apparent size of biopharmaceutical proteins. This underpins the appropriate use of such CE data in support of regulatory submissions.     

A Series of Collaborations between Various Pharmaceutical Companies and Regulatory Authorities Concerning the Analysis of Biomolecules Using Capillary Electrophoresis: Additional Instruments/Buffer

An international project team (including members from US, Canada and UK) was formed from a number of interested biopharmaceutical companies and regulatory authorities to conduct a cross-organisation collaboration exercise. The results of the first comparison with eight different organisations that used instruments of the same equipment model, the same reagents, and the same methodology has been reported previously [1]. This report represents the addition of other instruments using a different run buffer. The relative migration times were different, as expected, prohibiting a direct comparison between companies. The within-organisation variability was low for both relative migration time (<0.34% RSD% for all companies save one) and the peak area (<5% RSD% for all companies save one) when measuring the purity of a representative IgG sample. The apparent molecular weight of bovine serum albumin was measured with good precision (less than 10% RSD% across all companies) to the theoretical value when all data is utilized (67.5 kDa compared to 66.4 kDa). For a representative IgG sample, the three main components, IgG Light Chain, IgG Non-glycosylated Heavy Chain, and IgG Heavy Chain, could not be separated, specifically the IgG Non-glycosylated Heavy Chain and IgG Heavy Chain. When the IgG Non-glycosylated Heavy Chain and IgG Heavy Chain were combined for all organisations, the fractional peak area for the IgG Light Chain and IgG Non-glycosylated Heavy Chain + IgG Heavy Chain peak also showed excellent agreement, with less than 7.5 and 3.5% RSD%, respectively. The value of this exercise is in demonstrating the reliability of CE for the determination of apparent size of biopharmaceutical proteins. This underpins the appropriate use of such CE data in support of regulatory submissions.