NDLI: Advantages of CE-SDS over SDS-PAGE in mAb purity analysis

Content Provider	Royal Society of Chemistry (RSC)
Author	Li, Zhen Lin, Jun Shi, Ying
Copyright Year	2012
Abstract	Traditional SDS-PAGE and a newly developed CE-based method, i.e., CE-SDS, are both widely used to determine mAb purity. This work aimed to compare the two methods side-by-side in details and to establish the pros and cons of each. The sample preparation conditions were first optimized to acquire fair comparison results. When analyzing stressed mAb samples, both methods consistently showed comparable band/peak patterns. The results also suggest both methods are stability indicating. Beyond that, CE-SDS had improved advantages over SDS-PAGE in assay precision, linearity and separation resolution etc. Analysis of the same sample 21 times by utilizing SDS-PAGE introduced large standard deviation in LC and HC percentage determination. Furthermore, SDS-PAGE performed poorly in the linearity testing. The correlation coefficients from the regression were all less than 0.93 (SDS-PAGE) in contrast to those of greater than 0.97 (CE-SDS). When used to separate the non-glycosylated heavy chain (NGHC) from the heavy chain (HC), CE-SDS clearly showed much better resolution over SDS-PAGE. The CE-SDS method was further validated to assess its assay specificity, precision and robustness. For LC, HC and NGHC, the R.S.D. for repeatability, intraday and intermediate precision were determined to be <0.5% for migration time, <2.0% for corrected area percentage and <5.0% for corrected area, respectively. Finally, a small deviation in the migration time and corrected peak areas demonstrated the robustness of the CE-SDS assay when using different capillaries, analysts and sample storage conditions. All these results indicated that the newly developed CE-SDS method is superior to SDS-PAGE, and is suitable for use in areas of biochemical and biological research.
Starting Page	1637
Ending Page	1642
Page Count	6
File Format	HTM / HTML PDF
ISSN	17599660
Volume Number	4
Issue Number	6
Journal	Analytical Methods
DOI	10.1039/c2ay25208b
Language	English
Publisher	Royal Society of Chemistry
Access Restriction	Open
Subject Keyword	NGHC Polyacrylamide gel electrophoresis Standard deviation
Content Type	Text
Resource Type	Article
Subject	Analytical Chemistry Engineering Chemical Engineering

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