The development of capillary electrophoresis, especially CE‐SDS devices, has led CE‐SDS to become an established tool in a wide range of applications in the analysis of biopharmaceuticals and is increasingly replacing its method of origin, SDS‐PAGE. The goal of this study was to evaluate the comparability of molecular weight (MW) determination especially by CE‐SDS and SDS‐PAGE. For ensuring comparability, model proteins that have little or no posttranslational modifications and an IgG antibody were used. Only a minor influence of sample preparation conditions, including sample buffer, temperature conditions, and different reducing agents on the MW determination were found. In contrast, the selection of the MW marker plays a decisive role in determining the accurate apparent MW of a protein. When using different MW markers, the deviation in MW determination can exceed 10%. Interestingly, CE‐SDS and 10% SDS‐PAGE hardly differ in their trueness of MW determination. The trueness in relation to the reference MW for each protein was calculated. Although the trueness values for the model proteins considered range between 1.00 and 1.11 using CE‐SDS, they range between 0.93 and 1.03 on SDS‐PAGE, depending on the experimental conditions chosen.

中文翻译：

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CE-SDS、SDS-PAGE 和 Simple Western 的比较研究：样品制备对蛋白质分子量测定的影响

毛细管电泳，尤其是 CE-SDS 设备的发展，使 CE-SDS 成为生物药物分析中广泛应用的成熟工具，并越来越多地取代其起源方法 SDS-PAGE。本研究的目的是评估分子量 (MW) 测定的可比性，尤其是通过 CE-SDS 和 SDS-PAGE。为了确保可比性，使用了很少或没有翻译后修饰的模型蛋白质和 IgG 抗体。发现样品制备条件（包括样品缓冲液、温度条件和不同还原剂）对 MW 测定的影响很小。相比之下，MW 标记的选择在确定蛋白质的准确表观 MW 方面起着决定性作用。当使用不同的 MW 标记时，MW 测定的偏差可超过 10%。有趣的是，CE-SDS 和 10% SDS-PAGE 在 MW 测定的真实性上几乎没有区别。计算每种蛋白质相对于参考 MW 的真实性。尽管使用 CE-SDS 所考虑的模型蛋白质的真实度值介于 1.00 和 1.11 之间，但它们在 SDS-PAGE 上的范围介于 0.93 和 1.03 之间，具体取决于所选的实验条件。