A decade of microchip electrophoresis for clinical diagnostics – a review of 2008-2017

A core element in clinical diagnostics is the data interpretation obtained through the analysis of patient samples. To obtain relevant and reliable information, a methodological approach of sample preparation, separation, and detection is required. Traditionally, these steps are performed independently and stepwise. Microchip capillary electrophoresis (MCE) can provide rapid and high-resolution separation with the capability to integrate a streamlined and complete diagnostic workflow suitable for the point-of-care setting. Whilst standard clinical diagnostics methods normally require hours to days to retrieve specific patient data, MCE can reduce the time to minutes, hastening the delivery of treatment options for the patients. This review covers the advances in MCE for disease detection from 2008 to 2017. Miniaturised diagnostic approaches that required an electrophoretic separation step prior to the detection of the biological samples are reviewed. In the two main sections, the discussion is focused on the technical set-up used to suit MCE for disease detection and on the strategies that have been applied to study various diseases. Throughout these discussions MCE is compared to other techniques to create context of the potential and challenges of MCE. A comprehensive table categorised based on the studied disease using MCE is provided. We also comment on future challenges that remain to be addressed

Sensitivity enhancing injection from a sample reservoir and channel interface in microchip electrophoresis

The stacking of a cationic analyte (i.e., rhodamine B) at the interface between a sample reservoir and channel in a microchip electrophoresis device is described for the first time. Stacking at negative polarity was by micelle to solvent stacking where the dye was prepared in a micellar solution (5 mM sodium dodecyl sulfate in 25 mM phosphoric acid, pH 2.5) and the channel was filled with high methanol content background solution (70% methanol in 50 mM phosphoric acid, pH 2.5). The injection of the stacked dye into the channel was by simple reversal of the voltage polarity with the sample solution and background solution at the anodic and cathodic reservoirs of the straight channel, respectively. The enrichment of rhodamine B at the interface and injection of the stacked dye into the channel was clearly visualized using an inverted fluorescence microscope. A notable sensitivity enhancement factor of up to 150 was achieved after 2 min at 1 kV of micelle to solvent stacking. The proposed technique will be useful as a concentration step for analyte mixtures in simple and classical cross-channel microchip electrophoresis devices or for the controlled delivery of enriched reagents or analytes as narrow plugs in advanced microchip electrophoresis devices

Capillary electrophoresis: overview

In capillary electrophoresis (CE), analytes are separated under the influence of an electric field. The CE family of separation techniques is a complementary analytical technique to liquid chromatography, which are both used for quantitation and identification of target analytes. This overview outlines the instrumental parts involved in the electrophoretic separation process and briefly discusses the history of CE and its technological advancement. An introduction is provided to the fundamental parameters, separation capillary, capillary modifications, sample injection, stacking, and detection

Unusual stacking with electrokinetic injection of cationic analytes from micellar solutions in capillary zone electrophoresis

Electrokinetic injection (EKI) in capillary zone electrophoresis (CZE) of charged analytes is by the electroosmotic flow (EOF) and electrophoretic mobility of analytes. In most forms of stacking with EKI, the sample ions were introduced via electrophoretic mobility and concentrated in a stacking boundary inside the capillary. In this work, we describe the unusual stacking of cationic analytes via EKI of sodium dodecyl sulfate (SDS) micelles into a fused silica capillary filled with acidic background solution (BGS) with 40–50\ua0% acetonitrile. The analytes prepared with SDS micelles were injected because of their interaction with micelles or effective electrophoretic mobility. We observed two peaks from an analyte, and this suggested the concentration of analytes into two stacking zones. These two adjacent stacking zones were surprisingly maintained inside the capillary during EKI although the EOF was moving towards the inlet. The zones were identified as the SDS micelles (micelles zone) and organic solvent-rich stacking zone (solvent-rich zone) where the micelles zone was closer to the inlet end of capillary. The analytes concentrated in the solvent-rich zone through the mechanism of micelle to solvent stacking (MSS). The concentrated analytes in the micelles zone were from the concentrated analytes that electrophoretically migrated into the micelles zone from the solvent-rich zone during EKI. The analytes in the micelles zone were then re-stacked by MSS and formed the second sharp peak in CZE. This was prevented by reduction of acetonitrile concentration in the inlet BGS. A sensitivity enhancement factor of more than 100 was obtained for model cationic drugs (diphenhydramine and imipramine)

Derivatisation for separation and detection in capillary electrophoresis (2015–2017)

Derivatisation is an integrated part of many analytical workflows to enable separation and detection of the analytes. In CE, derivatisation is adapted in the four modes of pre-capillary, in-line, in-capillary, and post-capillary derivatisation. In this review, we discuss the progress in derivatisation from February 2015 to May 2017 from multiple points of view including sections about the derivatisation modes, derivatisation to improve the analyte separation and analyte detection. The advancements in derivatisation procedures, novel reagents, and applications are covered. A table summarising the 46 reviewed articles with information about analyte, sample, derivatisation route, CE method and method sensitivity is provided

Stacking or On-line Sample Concentration in CE-MS for Metabolomics

ESI-MS is a powerful/sensitive detector for CE in metabolomics. For trace analysis of certain metabolites, off-line sample preparation will be required prior to CE-ESI-MS. An alternative and complementary sample preparation approach is stacking, which was initially developed for CE with UV detection to improve detection sensitivity. Stacking also offers the possibility of sample clean-up during analyte focusing, providing the opportunity to improve the tedious and long analysis commonly associated with bioanalysis. This chapter introduces the different stacking techniques developed in CE-UV, and their evolution into CE-ESI-MS with special emphasis on applications for metabolomics. The nature of metabolites (typically charged small molecules) makes it easy to apply the different stacking techniques currently available in the literature. However, it seems like stacking in CE-ESI-MS for metabolomics is still in its development/testing stage, and hopefully younger scientists will pursue research in this relevant area of study

Derivatisation for separation and detection in capillary electrophoresis (2012-2015)

Derivatisation is a well-established and mature form of sample preparation for CE. The modification of the analyte can cause superior analysis characteristics such as better sensitivity and selectivity, however, derivatisation of the analyte introduces an additional step into the analytical workflow. This review covers articles from January 2012 to January 2015 on derivatisation in CE. The main sections are on the derivatisation modes (i.e. pre-capillary, in-line, in-capillary and post-capillary), separation and detection modes (i.e. LIF and others). LIF is discussed in more detail since this detection mode was most prevalent. A table of the common labelling agents and wavelengths for excitation and emission and the common derivatisation reactions are included. In addition, a comprehensive table which summarises all research articles is provided. This review is suitable for analytical chemists as a guide for 'how to get started' with derivatisation for separation and detection in CE

High Performance Liquid Chromatography versus Stacking-Micellar Electrokinetic Chromatography for the Determination of Potentially Toxic Alkenylbenzenes in Food Flavouring Ingredients

Alkenylbenzenes, including eugenol, methyleugenol, myristicin, safrole, and estragole, are potentially toxic phytochemicals, which are commonly found in foods. Occurrence data in foods depends on the quality of the analytical methodologies available. Here, we developed and compared modern reversed-phase high performance liquid chromatography (HPLC) and stacking-micellar electrokinetic chromatography (MEKC) methods for the determination of the above alkenylbenzenes in food flavouring ingredients. The analytical performance of HPLC was found better than the stacking-MEKC method. Compared to other HPLC methods found in the literature, our method was faster (total run time with conditioning of 15 min) and able to separate more alkenylbenzenes. In addition, the analytical methodology combining an optimized methanol extraction and proposed HPLC was then applied to actual food flavouring ingredients. This methodology should be applicable to actual food samples, and thus will be vital to future studies in the determination of alkenylbenzenes in food