Analytical methods electrophoresis

Boss H J., Watson D.B., and Rush R.S. (1998), Peptide capillary zone electrophoresis mass spectrometry of recombinant human erythropoietin an evaluation of the analytical method, Electrophoresis 19(15), 2654—2664. 

Juan HF, Lin JYC, Chang WH, Wu CY, Pan TL, Tseng MJ, Khoo KH, Chen ST. Biomic study of human meyloid leukemia cells differentiation to macrophages using DNA array, proteomic, and bioinformatic analytical methods. Electrophoresis 2002 23 (15) 2490-504. 

The use of agarose as an electrophoretic method is widespread (32—35). An example of its use is in the evaluation and typing of DNA both in forensics (see Forensic chemistry) and to study heritable diseases (36). Agarose electrophoresis is combined with other analytical tools such as Southern blotting, polymerase chain reaction, and fluorescence. The advantages of agarose electrophoresis are that it requires no additives or cross-linkers for polymerization, it is not hazardous, low concentration gels are relatively sturdy, it is inexpensive, and it can be combined with many other analytical methods. 

Two-Dimensional Electrophoresis. Two-dimensional (2D) electrophoresis is unique, offering an analytical method that is both reproducible and sensitive. It is referred to as 2D because it employs two different methods of electrophoresis, in two different dimensions, to produce one result. Each method separates the sample compounds based on different properties of each compound. The combination of the two methods gives better resolution of the compounds in the sample than could be achieved with either method alone. For example, each method alone may separate up to 100 components of a sample, whereas together they may separate up to 10,000 components. 

Identification of stmctures of toxic chemicals in environmental samples requires to use modern analytical methods, such as gas chromatography (GC) with element selective detectors (NPD, FPD, AED), capillary electrophoresis (CE) for screening purposes, gas chromatography/mass-spectrometry (GC/MS), gas chromatography / Fourier transform infra red spectrometry (GC/FTIR), nucleai magnetic resonance (NMR), etc. 

At present moment, no generally feasible method exists for the large-scale production of optically pure products. Although for the separation of virtually every racemic mixture an analytical method is available (gas chromatography, liquid chromatography or capillary electrophoresis), this is not the case for the separation of racemic mixtures on an industrial scale. The most widely applied method for the separation of racemic mixtures is diastereomeric salt crystallization [1]. However, this usually requires many steps, making the process complicated and inducing considerable losses of valuable product. In order to avoid the problems associated with diastereomeric salt crystallization, membrane-based processes may be considered as a viable alternative. 

The refinement of other analytical methods, such as electrophoresis [34,36], the various techniques of optical spectroscopy [103-105], and nuclear magnetic resonance [201], is supplemented by the recent advances in real-time affinity measurements [152,202], contributing to the understanding of biomolecular reactivity. Taken together, the improvement of analytical methods will eventually allow a comprehensive characterization of the structure, topology, and properties of the nucleic acid-based supramolecular components under consideration for distinctive applications in nanobiotechnology. 

Detection of the PSP toxins has proven to be one of the largest hurdles in the development of analytical methods. The traditional means, and still in wide use today, is determination of mouse death times for a 1 mL injection of the test solution. There are a variety of drawbacks to utilization of this technique in routine analytical methods, that have prompted the search for replacements. In 1975 Bates and Rapoport (3) reported the development of a fluorescence technique that has proven to be highly selective for the PSP toxins, and very sensitive for many of them. This detection technique has formed the basis for analytical methods involving TLC (77), electrophoresis (72), column chromatography (7J), autoanalyzers (7 ), and HPLC (5,6,7). 

The extraordinary complexity of human genes and their products has encouraged the development of extremely high-resolution analytical methods.75 Capillary electrophoresis is competitive with slab gel methods, with resolution up to the order of about 1,000 base pairs for sequencing, sizing, and detection of mutation. Reversed phase HPLC is useful for restriction digest mapping and MALDI-MS up to about 1000 base pairs. 

Capillary electrophoresis employing chiral selectors has been shown to be a useful analytical method to separate enantiomers. Conventionally, instrumental chiral separations have been achieved by gas chromatography and by high performance liquid chromatography.127 In recent years, there has been considerable activity in the separation and characterization of racemic pharmaceuticals by high performance capillary electrophoresis, with particular interest paid to using this technique in modem pharmaceutical analytical laboratories.128 130 The most frequently used chiral selectors in CE are cyclodextrins, crown ethers, chiral surfactants, bile acids, and protein-filled... 

There are a few other analytical methods in which electrochemistry plays an essential role, such as (paper) electrophoresis, isotachophoresis, electrography and electrochromatography (according to Fujinaga) as they belong to analytical separation techniques, they are beyond the scope of this book. 

Capillary electrophoresis has also been combined with other analytical methods like mass spectrometry, NMR, Raman, and infrared spectroscopy in order to combine the separation speed, high resolving power and minimum sample consumption of capillary electrophoresis with the selectivity and structural information provided by the other techniques [6]. 

Becker H., Gartner C., Polymer microfabrication methods for microfluidic analytical applications, Electrophoresis 2000 21 12-26. 

Microfluidics evolved from micro-analytical methods in capillary format such as capillary electrophoresis, high-performance liquid chromatography, and gas chromatography, and has successfully revolutionized chemical and biochemical... 

H. Becker and C. Gartner, Polymer microfabrication methods for microfludic analytical applications. Electrophoresis 21, 12—26 (2000). 

The principal analytical methods for complex samples are those that separate the mixture by differential migration and then detect the separated components. The separation methods are chromatography, electrophoresis, and field flow fractionation the detection methods—which need not be selective but must be sensitive—include absorption, laser-induced fluorescence, electrochemistry, and mass... 

In the last decade, capillary electrophoresis (CE) has become one of the most powerful and conceptually simple separation techniques for the analysis of complex mixtures. The main reasons are its high resolution, relatively short analysis times, and low operational cost when compared to high-performance liquid chromatography (HPLC). The ability to analyze ultrasmall volume samples in the picoliter-to-nanoliter ranges makes it an ideal analytical method for extremely volume-limited biological microenvironments. 

Application of new analytical methods, in particular of different types of electrophoresis, has also provided new data on the molecular properties of these enzymes and has set more rigorous criteria of their purity and homogeneity. 

Analytical methods are ripe for attack using Al methods. Capillary electrophoresis is a routine separation technique, but like other separation techniques, its effectiveness is correlated strongly with experimental conditions. Hence it is important to optimize experimental conditions to achieve the maximum degree of separation. Zhang and co-workers41 studied the separation of mixtures in reserpine tablets, in which vitamin B1 and dibazolum may be incompletely separated, as may promethazine hydrochloride and chloroquine... 

Gel electrophoresis, a common analytical method in biological and medical research, can be conveniently applied in the analysis of dendritic polymers, owing primarily to the unique structure of the dendrimer analytes. It has currently been used to assess purity, measure molecular weight of dendrimers,... 

Microdialysis is a sampling technique that must be coupled with an analytical method to identify and quantify chemical components of the dialysate. The samples can be analyzed immediately upon collection (i.e., online), or they can be stored (—80°C) for future analysis. Only analytical techniques sensitive enough to measure both small sample volumes and low concentrations of substances can be used to measure compounds in dialysate samples. High-performance liquid chromatography (HPLC) or capillary electrophoresis (CE) combined with ultraviolet (UV), electrochemical (EC), or laser-induced fluorescence (LIF)... 

Apart from the qualification dossiers provided by vendors there seems, at present, to be very little information published on the performance of an operational qualification for capillary electrophoresis (CE) instruments other than a chapter in Analytical Method Validation and Instrument Performance. The chapter, written by Nichole E. Baryla of Eli Lilly Canada, Inc, discusses the various functions (injection, separation, and detection) within the instrument and provides guidance on the type of tests, including suggested acceptance criteria, that may be performed to ensure the correct working of the instrument. These include injection reproducibility and linearity, temperature and voltage stability, detector accuracy, linearity, and noise. 

VALIDATION OF ANALYTICAL METHODS USING CAPILLARY ELECTROPHORESIS... 

---