Capillary electrophoresis instruments

Swedberg (1997), Fernandes and Flick (1998) and Frazier et al. (2000a) have described the principles and some applications of CE. A recent review by Kok (2000) describes capillary electrophoresis instrumentation and operation. The major formats of CE are ... 

In this chapter, it will be shown how to build the capillary electrophoresis instrument from its components into a robust, functioning analytical instrument. The various components are presented, including a wide range of detection techniques. Some basic troubleshooting along with advice on how to qualify the system is presented along with future prospects for the technique. 

Qualification of a capillary electrophoresis instrument is performed using failure mode, effects, and criticality analysis as the risk analysis tool. The instrument is broken down into its component modules and the potential failures of those components identified. The potential effect of those failures is defined and the risk characterized. Any current evaluation of those failures is identified and any recommended actions to mitigate the risk defined. 

A schematic diagram of a capillary electrophoresis instrument is shown in Figure... 

Figure 6.1 A schematic representation of the arrangement of the main components of the capillary electrophoresis instrument.

Francois, Y., Zhang, K., Varenne, A., and Gareil, R, New integrated measurement protocol using capillary electrophoresis instrumentation for the determination of viscosity, conductivity and absorbance of ionic liquid-molecular solvent mixtures. Anal. Chim. Acta, 562,164-170, 2006. 

For analytical equipment, qualification is broken down into four areas design qualification (DQ), installation qualification (IQ), operational qualification (OQ), and performance verification (PV) [2,3]. In this chapter we focus on the operational qualification of a capillary electrophoresis instrument. The tests used in the operational qualification are often used in the routine performance verification as... 

Figure 8.3—Schematic of a standard capillary electrophoresis instrument. The electrolyte is an aqueous ionic solution that has been filtered and degassed. It can contain several additives. There are several processes that can be used to introduce the sample into the capillary (cf. 8.4). The use of voltages above 30 kV is rare because they require special insulation. The length of the capillary (L) and the effective distance of migration (/) must not be confused since the latter is shorter by 10 or 20 cm.

In capillary electrophoresis instruments, the electro-osmotic flow is used to impose, on all charged species in the sample, a direction of migration that is oriented from the anode towards the cathode. An increase in the electro-osmotic flow vEOS decreases, at the detector, the gap in migration times of ions travelling in the same direction. The use of fused silica capillaries partially deactivated by coating the inner wall allows modulation of the electro-osmotic flow. A voltage gradient can also be used to this end. 

Nowadays, the demand for nanoanalyses is increasing continuously and some advancement has been made in chromatographic and capillary electrophoresis instruments. The integration of all the steps of analysis, that is, sample preparation, injection, separation, and detection on a single chip, is the most difficult task for scientists. In spite of many encouraging advancements in chip LC/CE instrumentation we are still far away from realizing the visions presented a decade ago. Briefly, more advances are needed to turn the dream of real nanochromatography and capillary electrophoresis into a mature analytical tool. 

The analysis of aliphatic acids was performed using a P/ACE MDQ capillary electrophoresis instrument equipped with a 60 cm x 50 pm id fused silica capillary (Beckman Coulter, Fullerton, CA). The samples were filtered through a 0.45-gm cellulose acetate filter (Whatman, Maidstone, UK) prior to hydrodynamic injection at 15 psi for 4 s. The voltage was set to 20 kV at reversed polarity. The electrolyte, composed of 5.0 mM trimellitic acid, 50 mM tris(hydroxymethyl)-aminomethane, 1.0 mM tetradecyl-trimethylammoniumbromide, and 0.5 mM calcium chloride, had a pH of 9.8. Before use, it was filtered through a 0.2-gm cellulose nitrate filter and degassed withhelium. Detection was performedby indirect UV absorption at 220 nm. Succinic acid was used as internal standard. 

One of the most important features of the capillary electrophoresis instrument is the detection system. Several of the detection methods commonly used in high-performance liquid chromatography have been somewhat adapted for capillary electrophoresis. Currently, the most popular detection methods developed for capillary electrophoresis are summarized in Table III. 

Despite the availability of several detectors that have been successfully adapted for capillary electrophoresis, only ultraviolet and noncoherent light fluorescence are currently configured in the capillary electrophoresis instruments. Presently, the most common way to enhance detection of... 

Another important component of the capillary electrophoresis instrumentation is the cassette-cartridge design (Figure 5). This modular component has three functions 1) protection of the capillary column, 2) the accomodation of capillary columns of... 

Kappes et al. evaluated the potentiometric detection of acetylcholine and other neurotransmitters through capillary electrophoresis [209]. Experiments were performed on an in-house capillary electrophoresis instrument that made use of detection at a platinum wire, dip-coated in 3.4% potassium tetrakis (4-chlorophenyl) borate/64.4% o-nitrohenyl octyl ether/32.2% PVC in THF. The results were compared to those obtained using capillary electrophoresis with amperometric detection at a graphite electrode. Samples prepared in the capillary electrophoresis buffer were electrokinetically injected (7 s at 5 kV) into an untreated fused silica capillary (88 cm x 25 pm i.d.) and separated with 20mM tartaric acid adjusted to pH 3 with MgO as the running buffer. The system used an applied potential of 30 kV, and detection versus the capillary electrophoresis ground electrode. 

Capillary electrophoresis is an exciting, new, high resolution separation technique useful for the determination of drugs and their metabolites in body fluids. The first commercial capillary electrophoresis instruments began to emerge on the market in 1988. Today approximately a dozen companies manufacture electrokinetic capillary instrumentation, with many of these fully automated, that comprise auto samplers with computerized data evaluation.f Capillary electrophoresis involves the electrophoretic separations of minute quantities of molecules in solution according to their different velocities in an applied electrical field. The velocity of these molecules... 

Capillary electrophoresis instrumentation is fully automated with respect to sample injection, separation, on-capillary detection, and postrun data analysis. 

Kok WT (2000) Capillary electrophoresis instrumentation and operation, Chro-matographia supplement, vol 51. Vieweg, Braunschweig... 

The OQ/PV should be performed on installation, after extended use (e.g. at least once per year) and after major repair. This may also be extended to performing an OQ/PV before starting to validate or optimise a method on that instrumentation. OQ/PV for a capillary electrophoresis instrument should therefore be directed towards those specific instrumental functions which can affect the CE analysis. 

From the perspective of the clinical laboratory, miniaturization has been a long-term trend in clinical diagnostics instrumentation. For example, capillary electrophoresis instruments (see Chapter 5) and mass spectrometers have been implemented on microchips of silicon, glass, or plastic. In actuality, however, these devices are not manufactured on a nanometer scale but rather on a micrometer scale. Consequently, this chapter will be concerned with microminiaturized devices whose key components (1) are approximately 100 micrometers in size, (2) are employed in analytical measurement, and (3) require special forms of fabrication designed for microdevices. Although this chapter does not attempt to discuss submicron or molecular structures at the nanometer scale, it should be noted that applications discussed later in it require only nanoliter (nL) quantities of a sample or deal with individual cells that may have cell volume in the picoHter (pL) to nL range. 

T Wehr, M Zhu, R Rodriguez, D Burke, K Duncan. High performance isoelectric focusing using capillary electrophoresis instrumentation. Am Biotech Lab. 8 22-29 1990. 

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