Capillary electrophoresis as a clinical tool

Clinical laboratories must produce accurate results for patients with a minimum turn-around time. Automated commercial capillary electrophoresis instrumentation has been available to the clinical laboratory for the past five years. Our laboratory has utilised capillary electrophoresis (CE) to automate serum protein electrophoresis. We have used the technique of CE to produce clinical results for nearly two years. CE methods are also available for the quantitation of haemoglobin variants, by both isoelectric focusing and free solution techniques. Micellar electrokinetic separations by CE have been developed for some specialised drug assays and for B-group vitamin analysis, while gel-filled capillaries have the capability to separate DNA fragments, such as PCR products. Isoenzyme analysis has shown possibilities by CE, but quantitative results are needed to be clinically useful. Analysis of amino acids for newborn screening programs and as an arterial clotting indicator are being developed. The next five years should see a proliferation of clinical laboratory methods using automated CE.     

Use of nanomaterials in capillary and microchip electrophoresis

This review gives an overview of different separation strategies with nanomaterials and their use in capillary electrophoresis (CE) and capillary electrochromatography, as well as in microchip electrophoresis, including metal and metal oxide nanoparticles, carbon nanotubes, fullerene and polymer nanoparticles, as well as silica nanoparticles. The paper highlights the new developments and innovative applications of nanoparticles as pseudostationary phases or immobilized on the capillary surface for CE separation. The separation and characterization of target nanoparticles with different sizes by CE are reviewed likewise.     

Capillary electrophoresis procedures for serum protein analysis: comparison with established techniques

Methods using automated capillary electrophoresis (CE) instrumentation are available for serum protein electrophoresis with monoclonal band quantitation, isoelectric focusing and sodium dodecyl sulphate-polyacrylamide gel electrophoresis separations. The advantages of CE over previous gel methods relate to the time and labour saved by the automated instrumentation. High pI monoclonal bands and cryoglobulin specimens can be successfully analysed by CE. However, if the CE application uses a standard company supplied kit, then the cost savings are often negated by the high cost of the kit. Improvements such as the inclusion of both a UV-Vis as well as a fluorescence detector as standard within the one commercial instrument, the production of coated IEF capillaries with a useful life of at least 100 samples, and the introduction of a capillary array into all commercial instrumentation would ensure greater use of CE within both the clinical and other protein laboratories.     

Enantioselective separations using capillary electrophoresis

The preconditions are outlined for enantioselective separations in capillary electrophoresis (CE) with chiral selectors as additives to the background electrolyte. Free solution capillary electrophoresis conditions are characterised by a single solution phase. Chiral separations are reviewed by selector type (chiral ligand exchange, cyclodextrins, crown ethers, glycoproteins) with the extensive studies on cyclodextrins grouped into sections on amino acids, pharmaceuticals, and speciality chemicals, optimisation, biological fluids, and quantitative aspects. In micellar electrokinetic capillary chromatography, enantioselective discrimination occurs by partition in a two-phase system, with a chiral micellar phase as selector. Optimum separation conditions can be readily predicted for a given selector–selectand combination, and absolute values of binding constants determined by CE. Advantages of CE in comparison with HPLC using a chiral stationary phase include robust, rapid assays and the use of small volumes of aqueous solutions; disadvantages include less favourable detection limits. © 1994 Wiley-Liss, Inc.     

Miniaturized electrophoresis: an evolving role in laboratory medicine.

The promise of capillary electrophoresis (CE) for supplanting conventional methods in the clinical laboratory led to intense interest in this analytical tool a decade ago. Since then, a number of clinical applications have been defined along with those that have impacted the pharmaceutical, environmental, and forensic arenas. Concurrent with the development of CE applications was the emergence of electrophoresis in the microchip format. The main attraction of this platform, the ability to execute high-resolution separations in a few hundred seconds, was not its only attribute. The capability for parallel processing of separations was complemented by the potentialfor integrating sample preparation into the same device. This Review highlights recent progress towards CE and microchip electrophoresis as clinical diagnostic tools, with literature coverage from 1996 to 2000.     

Determination of illicit and/or abused drugs and compounds of forensic interest in biosamples by capillary electrophoretic/electrokinetic methods

The application of capillary electrophoresis (CE) methods in forensic toxicology for the determination of illicit and/or misused drugs in biological samples is reviewed in the present paper. Sample pretreatments and direct injection modes used in CE for analysis of drugs in biological fluids are briefly described. Besides, applications of separation methods based on capillary zone electrophoresis or micellar electrokinetic chromatography with UV absorbance detection to (i) analysis of drugs of abuse, (ii) analysis of other drugs and toxicants of potential forensic interest and (iii) for metabolism studies are reviewed. Also, alternative CE methods are briefly discussed, including capillary isotachophoresis and separation on mixed polymer networks. High sensitivity detection methods used for forensic drug analysis in biological samples are then presented, particularly those based on laser induced fluorescence. A glimpse of the first examples of application of CE–mass spectrometry in forensic toxicology is finally given.     

Large-volume sample stacking of selected drugs of forensic significance by capillary electrophoresis

Large-volume sample stacking capillary electrophoresis (LVSS-CE) and conventional capillary electrophoresis (CE) are compared for the separation of drugs of significance to forensic and clinical analyses. LVSS-CE for cations requires the use of an electroosmotic flow (EOF) modifier in conjunction with polarity switching to effect on-column concentration of an analyte and its subsequent migration in the capillary. The run buffer consists of 0.05 mol dm⁻³ disodium tetraborate adjusted to pH 2.2 with orthophosphoric acid, and the EOF modifier is 0.002 mol dm⁻³ cetyltrimethylammonium bromide. CE investigations used an identical run buffer minus the EOF modifier. LVSS-CE and CE investigations used injection times of 30 s and 3 s, respectively. Both modes of capillary electrophoresis are compared in terms of their limits of detection, efficiency, resolution and reproducibility. LVSS-CE is also applied to the analysis of a spiked urine sample.     

Recent developments of enantioseparation techniques for adrenergic drugs using liquid chromatography and capillary electrophoresis: a review

This review provides the achievements of enantioseparation of adrenergic drugs and application of these methods in clinical and pharmaceutical analysis. The adrenergic agonist and antagonist drugs are analyzed in the direct and indirect modes by liquid chromatography (LC) and capillary electrophoresis (CE). Other chromatographic enantioseparation methods including super- and sub-critical fluid chromatography (SFC), and capillary electrochromatography (CEC) are presented likewise to analyse the cited compounds. The different separation processes for these drugs are briefly discussed and some applications are presented.     

Characterization of proteins in the human serum proteome.

This paper presents a multidimensional profile of the human serum proteome, produced by a two-dimensional protein fractionation system based on liquid chromatography followed by characterization with capillary electrophoresis (CE). The first-dimension separation was done by chromatofocusing over a pH range from 8.5 to 4.0, where proteins were separated by their isoelectric points (pI). In this dimension, fractions were collected based on pH. The first-dimension pI fractions were then resolved in the second dimension by high-resolution, reversed-phase chromatography with a gradient of trifluoroacetic acid (TFA) in acetonitrile and TFA in water. A selected protein fraction collected from the second dimension by time was characterized by CE for molecular-weight estimation and for presence of isoforms. Molecular-weight estimation was done by sodium dodecyl sulfate capillary gel electrophoresis, where proteins were separated in the range of 10,000-225,000 Da. Detection of isoforms was done by capillary isoelectric focusing over a pH range of 3-10. A selected second-dimension fraction that contained the putative serum iron-binding protein transferrin was analyzed by these two CE techniques for molecular-weight determination and the presence of isoforms. The combination of two-dimensional protein fractionation and CE characterization represents an advanced tool for proteomics.     

Use of nanomaterials in capillary and microchip electrophoresis

This review gives an overview of different separation strategies with nanomaterials and their use in capillary electrophoresis (CE) and capillary electrochromatography, as well as in microchip electrophoresis, including metal and metal oxide nanoparticles, carbon nanotubes, fullerene and polymer nanoparticles, as well as silica nanoparticles. The paper highlights the new developments and innovative applications of nanoparticles as pseudostationary phases or immobilized on the capillary surface for CE separation. The separation and characterization of target nanoparticles with different sizes by CE are reviewed likewise.     

毛细管电泳-质谱联用技术研究进展

毛细管电泳-质谱（CE-MS）联用技术兼具毛细管电泳高效分离能力与质谱高灵敏检测、高真度定性的优势,已成为物质分离分析研究的一种非常重要的工具。本文对近几年来 CE-MS 联用的关键技术及 CE-MS 在中药分析、环境检测等领域的一些应用进展进行综述,对其发展进行了展望。     

Separation methods for pharmacologically active xanthones

Xanthones, as a kind of polyphenolic natural products with many strong bioactivities, are attractive for separation scientists due to the similarity and diversity of their structures resulting in difficult separation by chromatographic methods. High performance liquid chromatography (HPLC) and thin layer chromatography (TLC) are traditional methods to separate xanthones. Recently, capillary electrophoresis (CE), as a micro-column technique driven by electroosmotic flow (EOF), with its high efficiency and high-speed separation, has been employed to separate xanthones and determine their physicochemical properties such as binding constants with cyclodextrin (CD) and ionization constants. Since xanthones have been used in clinic treatment, the development of chromatographic and CE methods for the separation and determination of xanthones plays an essential role in the quality control of some herbal medicines containing xanthones. This article reviewed the separation of xanthones by HPLC, TLC and CE, citing 72 literatures. This review focused on the CE separation for xanthones due to its unique advantages compared to chromatographic methods. The comparison of separation selectivity of different CE modes including capillary zone electrophoresis (CZE), micellar electrokinetic chromatography (MEKC), microemulsion electrokinetic capillary chromatography (MEEKC) and capillary electrochromatography (CEC) was discussed. Compared with traditional chromatographic methods such as HPLC and TLC, CE has higher separation efficiency, faster separation, lower cost and more flexible modes. However, because of low sensitivity of UV detector and low contents of xanthones in herbal medicines, CE methods have seldom been applied to the analysis of real samples although CE showed great potential for xanthone separation. The determination of xanthones in herbal medicines has been often achieved by HPLC. Hence, how to enhance CE detection sensitivity for real sample analysis, e.g. by on-line preconcentration and CE-MS, would be a key to achieve the quantitation of xanthones.     

High-throughput single-strand conformation polymorphism analysis by capillary electrophoresis

Mutation detection plays a great role in genetic and medical research and clinical diagnosis of inherited diseases and particular cancers. Single-strand conformation polymorphism (SSCP) analysis is one of the most popular methods for detection of mutations. Recently, automated capillary electrophoresis (CE) systems have been used in SSCP analysis instead of conventional slab gel electrophoresis. SSCP analysis in combination with CE is a rapid, simple, sensitive and high-throughput mutation screening tool, and has been successfully applied for mutation detection involving human tumor suppressor genes, oncogenes and disease-causing genes. The new technique has a great potential for mutation screening of large numbers of samples in clinical diagnosis. This review discusses basic issues about the methodology of SSCP analysis based on CE and summarizes several key applications.     

Capillary electrophoresis-mass spectrometry as a characterization tool for therapeutic proteins

With the increasing use of capillary electrophoresis (CE) in the biotechnology industry, there is a demand for analytical tools and methodology that can be used to characterize CE profiles. This article describes the implementation and optimization of a robust online CE-mass spectrometry (CE-MS) system used for the characterization of several CE assays developed at Genentech Inc. These assays include CE as a complement to reverse-phase peptide mapping for the identification of small peptides eluting in the void volume, profiling N-linked glycopeptide heterogeneity, and determining O-linked site occupancy. In addition, CE-MS was used to confirm major 8-aminopyrene-1,3,6-trisulfonate (APTS)-labeled glycans released from recombinant antibodies that are routinely profiled by CE-laser-induced fluorescence (CE-LIF). For each study, CE-MS was able to successfully identify components seen in UV or LIF electropherograms, thereby expanding the capability of CE and CE-MS for profiling biomolecules.     

毛细管电泳及其在蛋白质折叠研究中的应用

简要介绍了毛细管电泳的常用分离模式及其原理,并对毛细管电泳在蛋白质化学领域中的新应用——研究蛋白质折叠和发展前景作了评述。     

Capillary electrophoresis

While capillary electrophoresis, or historically related techniques, have been used for over a century, and recognition of the value of this separation methodology has certainly grown rapidly in the past few years, the technique has generally been used by analytical chemists, particularly in Europe and Japan, and small groups of researchers in the United States. Many of the basic instrumentation problems have been solved only relatively recently, and researchers using capillary electrophoresis are now turning their attention to studying specific applications which demonstrate the potential versatility of this electrophoretic technique. The appearance of standardized commercial instrumentation is imminent. With the availability of such technology, capillary electrophoresis will no longer be an academic curiosity, but rather a tool with the potential for routine separations of diverse samples of interest to analyst, researcher, and clinician.     

Analysis of optical purity and impurity of synthetic D-phenylalanine products using sulfated beta-cyclodextrin as chiral selector by reversed-polarity capillary electrophoresis

A new capillary electrophoresis (CE) method has been achieved for simultaneous separation and quantification of phenylalanine, N-acetylphenylalanine enantiomers, and prochiral N-acetylaminocinnamic acid, possibly co-existent in reaction systems or synthesized products of D-phenylalanine. The separation was carried out in an uncoated capillary under reversed-electrophoretic mode. Among the diverse charged cyclodextrins (CDs) examined, highly sulfated (HS)-beta-CD as the chiral selector exhibited the best enantioselectivity. The complete separation of the analytes was obtained under the optimum conditions of pH 2.5, 35 mM Tris buffer containing 4% HS-beta-CD, applied voltage -15 kV, and capillary temperature 25 degrees C. Furthermore, the proposed method was applied to the determination of optical purity and trace impurities in three batches of the asymmetric synthetic samples of D-phenylalanine, and satisfactory results were obtained. The determination recoveries of the samples were in the range of 97.8-103.8%, and precisions fell within 2.3-5.0% (RSD). The results demonstrate that this CE method is a useful, simple technique and is applicable to purity assays of D-phenylalanine.     

Measurement of Free Choline in Plant Leaves by Capillary Electrophoresis

A low pH capillary electrophoresis (CE) was used for the measurement of free choline in plant leaves. Choline in the leaf extract was first converted to the benzoyl ester and put into CE. A well-resolved peak in the electropherogram was easily obtained. Involvement of enzymes in a two-step oxidation of choline to glycine betaine was evaluated in different plant species with the same mehod developed for glycine betaine and betaine aldehyde.     

Recent progress in capillary electrophoretic analysis of amino acid enantiomers

This review highlights recent progresses in capillary electrophoresis (CE) analysis of amino acid enantiomers in the last decade. Various chiral selectors including cyclodextrins (CDs), bile salts, crown ethers, cinchona alkaloids, metal-chiral amino acid complexes, macrocyclic antibiotics and proteins have been employed to separate amino acid enantiomers. In the CE analysis of amino acids, the selection of the separation mode is one of the most important issues to obtain good resolution of target enantiomers. Among several separation modes, CD-modified capillary zone electrophoresis (CD-CZE), CD electrokinetic chromatography (CDEKC), micellar EKC (MEKC), CD-modified micellar electrokinetic chromatography (CD-MEKC), capillary electrochromatography (CEC), ligand-exchange CE (LE-CE), and nonaqueous CE (NACE) have been employed to the chiral analysis of amino acids. More than 160 published research articles collected from SciFinder Scholar databases from the year 2001 described the enantioseparations of amino acids by capillary-based electrophoresis. This review provides a comprehensive table listing the CE analysis of amino acid enantiomers with categorizing by the separation modes.