Comparison of glycoprotein separation reveals greater impact of carbohydrates and disulfides on electrophoretic mobility for CE-SDS versus SDS-PAGE

Wang, Andrea L.; Paciolla, Mark; Palmieri, Michael J.; Hao, Gang

doi:10.1016/j.jpba.2019.113006

Cited by 10 publications

(4 citation statements)

References 25 publications

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“…The performance of slab gel versus capillary gels is similar (50), with only minor differences reported in mass range (50). Notably, the effect of glycosylation on molecular size determinations is more pronounced for capillary separations that incorporate linear sieving gels (51,52). The performance of multiple capillary instruments revealed no significant differences in assay performance and accuracy (53).…”

Section: Proteinsmentioning

confidence: 89%

Gels in Microscale Electrophoresis

Holland

Casto-Boggess

2023

Annual Rev. Anal. Chem.

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Gel matrices are fundamental to electrophoresis analyses of biopolymers in microscale channels. Both capillary gel and microchannel gel electrophoresis systems have produced fundamental advances in the scientific community. These analytical techniques remain as foundational tools in bioanalytical chemistry and are indispensable in the field of biotherapeutics. This review summarizes the current state of gels in microscale channels and provides a brief description of electrophoretic transport in gels. In addition to the discussion of traditional polymers, several nontraditional gels are introduced. Advances in gel matrices highlighted include selective polymers modified to contain added functionality as well as thermally responsive gels formed through self-assembly. This review discusses cutting-edge applications to challenging areas of discovery in DNA, RNA, protein, and glycan analyses. Finally, emerging techniques that result in multifunctional assays for real-time biochemical processing in capillary and three-dimensional channels are identified.

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Section: Proteinsmentioning

confidence: 89%

Gels in Microscale Electrophoresis

Holland

Casto-Boggess

2023

Annual Rev. Anal. Chem.

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“…Glycans could have a significant effect on safety and efficacy due to their effect on the folding and shielding of functional epitopes [71,72]. Glycans are known to affect the CE-SDS separation [73,74] and CE-SDS was used for glycoprotein analysis previously [38]. Therefore, the optimized CE-SDS method to determine the mini-HA protein by its primary structure required an optimal sample preparation protocol to reduce and deglycosylate the protein.…”

Section: Mini-ha Ptm Heterogeneitymentioning

confidence: 99%

Sixteen capillary electrophoresis applications for viral vaccine analysis

et al. 2021

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A broad range of CE applications from our organization is reviewed to give a flavor of the use of CE within the field of vaccine analyses. Applicability of CE for viral vaccine characterization, and release and stability testing of seasonal influenza virosomal vaccines, universal subunit influenza vaccines, Sabin inactivated polio vaccines (sIPV), and adenovirus vector vaccines were demonstrated. Diverse CZE, CE‐SDS, CGE, and cIEF methods were developed, validated, and applied for virus, protein, posttranslational modifications, DNA, and excipient concentration determinations, as well as for the integrity and composition verifications, and identity testing (e.g., CZE for intact virus particles, CE‐SDS application for hemagglutinin quantification and influenza strain identification, chloride or bromide determination in process samples). Results were supported by other methods such as RP‐HPLC, dynamic light scattering (DLS), and zeta potential measurements. Overall, 16 CE methods are presented that were developed and applied, comprising six adenovirus methods, five viral protein methods, and methods for antibodies determination of glycans, host cell‐DNA, excipient chloride, and process impurity bromide. These methods were applied to support in‐process control, release, stability, process‐ and product characterization and development, and critical reagent testing. Thirteen methods were validated. Intact virus particles were analyzed at concentrations as low as 0.8 pmol/L. Overall, CE took viral vaccine testing beyond what was previously possible, improved process and product understanding, and, in total, safety, efficacy, and quality.

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“…PAGE and CE-SDS. 83 They studied different mammalian glycoproteins utilizing these two tools under reduced and nonreduced conditions.…”

Section: Wang Et Al Compared Systematically Glycoprotein Migration On...mentioning

confidence: 99%

“…Five ribonuclease B glycoforms were resolved using CE in less than 20 min and the profile was in accord with that obtained using MALDI‐TOF‐MS. Wang et al compared systematically glycoprotein migration on SDS‐PAGE and CE‐SDS 83 . They studied different mammalian glycoproteins utilizing these two tools under reduced and nonreduced conditions.…”

Section: Experimental Toolsmentioning

confidence: 99%

Challenges and limitations in the studies of glycoproteins: A computational chemist's perspective

et al. 2021

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Experimenters face challenges and limitations while analyzing glycoproteins due to their high flexibility, stereochemistry, anisotropic effects, and hydration phenomena.Computational studies complement experiments and have been used in characterization of the structural properties of glycoproteins. However, recent investigations revealed that computational studies face significant challenges as well. Here, we introduce and discuss some of these challenges and weaknesses in the investigations of glycoproteins. We also present requirements of future developments in computational biochemistry and computational biology areas that could be necessary for providing more accurate structural property analyses of glycoproteins using computational tools. Further theoretical strategies that need to be and can be developed are discussed herein.

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Comparison of glycoprotein separation reveals greater impact of carbohydrates and disulfides on electrophoretic mobility for CE-SDS versus SDS-PAGE

Cited by 10 publications

References 25 publications

Gels in Microscale Electrophoresis

Gels in Microscale Electrophoresis

Sixteen capillary electrophoresis applications for viral vaccine analysis

Challenges and limitations in the studies of glycoproteins: A computational chemist's perspective

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