Investigation of an Uncommon Artifact during Reducing Capillary Electrophoresis-Sodium Dodecyl Sulfate Analysis of a Monoclonal Antibody with Dynamic Light Scattering and Reversed Phase High-Performance Liquid Chromatography

Han, Gao Rong; Wang, Si-Tao; Feng, Hao; Shen, Bin-Bin; Sun, Min-Fei; Wang, Haibin; Li, Lei; Fang, Wei-Jie

doi:10.1007/s11095-022-03303-0

Cited by 4 publications

(3 citation statements)

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“…Generally, CE-SDS analysis of monoclonal antibodies (mAbs) can be performed under reducing or nonreducing conditions. In reducing CE-SDS, the samples can be incubated with a reducing agent (e.g., β-mercaptoethanol (BME)), which can break the disulfide bond between the light chain (LC) and heavy chain (HC) [14][15][16]. LC, HC, and nonglycosylated HC (NGHC) can be separated from each other to help evaluate the purity of the sample under reducing conditions [13,14].…”

Section: Introductionmentioning

confidence: 99%

“…Similarly, in SDS-PAGE analysis of some proteins, artifacts were also caused by SDS binding restriction [24,25]. These problems all illustrated the limitations of SDS as a CE detergent, and the reasons for these problems are obviously related to the respective structures of different protein samples [16,26,27]. Due to the complexity of biological products, differences in specific amino acid sequences may cause huge differences in the chemical and physical properties of the samples, leading to inaccurate analysis, which also puts forward higher requirements for the analysis technique.…”

Section: Introductionmentioning

confidence: 99%

“…Finally, the use of the longer alkyl chain anionic surfactants sodium tetradecyl sulfate (STS) and sodium hexadecyl sulfate (SHS) as alternatives to SDS achieved the best optimization effect. SHS and STS have been previously reported for the optimization of HC or intact mAbs [16,22,23], but they have never been applied to improve LC peak performance as reported in this article. Here, we proved that SHS was also suitable for eliminating the tailing peak caused by the limitation of the complexation of SDS and LC in reducing CE-SDS, thereby improving the peak shape and providing a more accurate and stable analysis method.…”

Section: Introductionmentioning

confidence: 99%

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Elimination of light chain tailing in reducing capillary electrophoresis with sodium dodecyl sulfate analysis of a monoclonal antibody

et al. 2022

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Capillary electrophoresis with sodium dodecyl sulfate (CE‐SDS) is a common analytical technique for investigating the purity and molecular size heterogeneity of monoclonal antibody (mAb) drugs. In reducing CE‐SDS analysis of mAb‐A, the light chain (LC) peak exhibited severe tailing, seriously affecting the purity analysis. The purposes of this investigation are to clarify the source of tailing and develop a more appropriate CE‐SDS method to eliminate LC tailing. The degree of LC tailing was closely related to the mAb concentration, SDS concentration, and injection amount, and more hydrophobic detergents, such as sodium hexadecyl sulfate (SHS) and sodium tetradecyl sulfate (STS), could be used instead of SDS to obtain better peak shapes. The results also indicated that the tailing was caused by the binding problem associated with SDS, and SHS/STS could provide a more stable and uniform complexation for the LC. In summary, the method we developed successfully eliminated the LC tailing and provided a robust characterization of mAb‐A in reducing CE‐SDS analysis.

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