One-Step Focusing of Ampholytes With Electrophoretic Mobilization: Concepts Assessed by Computer Simulation.

IF 3 3区生物学 Q2 BIOCHEMICAL RESEARCH METHODS

ELECTROPHORESIS Pub Date : 2024-12-06 DOI:10.1002/elps.202400215

Wolfgang Thormann, Richard A Mosher

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Abstract

The dynamics of three one-step focusing protocols described in the literature for IEF-MS analyses of proteins are assessed by computer simulation. Focusing of 101 carrier ampholytes (pI range 3.0-11.0) and 9 analytes (pI range 3.9-10.4) with concurrent electrophoretic mobilization in an electroosmosis-free environment is studied via use of the following: (1) an acid as anolyte and catholyte, (2) a plug of a base between the sample and the acid as catholyte, and (3) a base incorporated into the acid-based catholyte. The data reveal that the first approach is of limited applicability and separation efficiency as it provides separated transient foci of acidic ampholytes only. The second system largely extends the applicability range, separation, and sensitivity and can be applied to ampholytes with pIs as high as 10. The same is true with the base incorporated into the catholyte. Dynamic simulation provides for the first time insight into the complexity of these one-step focusing systems with buffer components that are compatible with MS detection. Simulation is an attractive and simple tool for characterization of the involved electrophoretic processes and for assay optimization.

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来源期刊

ELECTROPHORESIS 生物-分析化学

CiteScore

6.30

自引率

13.80%

发文量

244

审稿时长

1.9 months

期刊介绍： ELECTROPHORESIS is an international journal that publishes original manuscripts on all aspects of electrophoresis, and liquid phase separations (e.g., HPLC, micro- and nano-LC, UHPLC, micro- and nano-fluidics, liquid-phase micro-extractions, etc.). Topics include new or improved analytical and preparative methods, sample preparation, development of theory, and innovative applications of electrophoretic and liquid phase separations methods in the study of nucleic acids, proteins, carbohydrates natural products, pharmaceuticals, food analysis, environmental species and other compounds of importance to the life sciences. Papers in the areas of microfluidics and proteomics, which are not limited to electrophoresis-based methods, will also be accepted for publication. Contributions focused on hyphenated and omics techniques are also of interest. Proteomics is within the scope, if related to its fundamentals and new technical approaches. Proteomics applications are only considered in particular cases. Papers describing the application of standard electrophoretic methods will not be considered. Papers on nanoanalysis intended for publication in ELECTROPHORESIS should focus on one or more of the following topics: • Nanoscale electrokinetics and phenomena related to electric double layer and/or confinement in nano-sized geometry • Single cell and subcellular analysis • Nanosensors and ultrasensitive detection aspects (e.g., involving quantum dots, "nanoelectrodes" or nanospray MS) • Nanoscale/nanopore DNA sequencing (next generation sequencing) • Micro- and nanoscale sample preparation • Nanoparticles and cells analyses by dielectrophoresis • Separation-based analysis using nanoparticles, nanotubes and nanowires.