The methodological developments in the field of capillary isoelectric focusing (CIEF) published between 1997-2001 are reviewed as a continuation of the previous review by Rodriguez-Diaz et al. (Electrophoresis 1997, 18, 2134-2144). The applications are summarized and the progress in CIEF technologies, including experimental setup with coated and uncoated capillaries, remedies for the presence of salts in samples, additives to reduce precipitation of samples during the focusing process, calibration of the pH gradients, issues of reproducibility, carrier ampholyte-free CIEF, and a computer simulation of focusing process are discussed. Developments of IEF separations in fabricated microchannels and the advances in detection schemes, i.e., imaging, fluorescence and chemiluminescence, are summarized. The progress in micropreparation was noted, and the massive works for two-dimensional separations are described for the coupling with size-exclusion chromatography and mass spectrometry, in which the developments aimed at proteomics are discussed separately. The applications for the detection of noncovalent complexes and the separations of microorganisms are reviewed.
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http://dx.doi.org/10.1002/elps.200290005 | DOI Listing |
Anal Biochem
January 2025
Advanced Electrophoresis Solutions Ltd., 380 Jamieson Parkway, Unit 7 and 8, ON, N3C 4N4, Canada; AES Biotech Jiaxing Ltd., No. 501 South Changsheng Road, Economic and Technological Development Zone, Jiaxing City, Zhejiang Province, PR China. Electronic address:
Characterizing major bovine milk proteins, including whey and casein, is of significant interest in the dairy industry. The diverse array of protein proteoforms can be different in terms of genetic variation, breed ways, lactation stage, and animal nutritional status. Current routine methods for bovine milk protein profiling are typically based on immunological techniques, infrared spectroscopy, slab gel isoelectric focusing, capillary electrophoresis, and high-performance liquid chromatography.
View Article and Find Full Text PDFAnal Chem
January 2025
Department of Chemistry and Biochemistry, University of Notre Dame, Notre Dame, Indiana 46556, United States.
Intact protein analysis using mass spectrometry (MS) is an important technique to characterize and provide a comprehensive overview of protein complexity. It is also the basis of "top-down" approaches in proteomics to describe the proteoforms of single protein's post-translational modifications (PTMs). MS-based analysis of intact proteins benefits from high-resolution separations prior to electrospray ionization.
View Article and Find Full Text PDFJ Sep Sci
January 2025
Department of Biosciences and Medical Biology, University of Salzburg, Salzburg, Austria.
Imaged capillary isoelectric focusing was successfully applied for separating an in-house synthesized closely related peptide pair, that is, a linear 12-mer (Rp5-L) and its cyclic 15-mer variant (Rp5-C). Rp5-L represents a mimotope, that is, an epitope mimicking peptide, of the CD20 antigen, which is over-expressed in B-cell-related tumors. Peptide identity-including the successful disulfide bond formation in Rp5-C-was confirmed with matrix-assisted laser desorption ionization-time of flight mass spectrometry.
View Article and Find Full Text PDFIntroduction: Cyanobacterium Arthrospira platensis (AP) (Nordstedt) Gomont contains high content of phycobiliproteins (PBP), which are an important source for food industry. Methods effectively extracting proteins contained in AP cells are demanded to provide a supply of the material. Water-based extraction methods are advisable due to the high solubility of the PBP.
View Article and Find Full Text PDFElectrophoresis
December 2024
RAM Software Solutions, Tucson, Arizona, USA.
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.
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