Electrophoretic separations background

Rawjee, Y. Y and Vigh, Gy., A peak resolution model for the capillary electrophoretic separation of the enantiomers of weak acids with hydroxypropyl (3-cyclodextrm-containing background electrolytes, Anal. Chem., 66, 619, 1994. 

We begin with a short introduction to provide polymer chemists who may be new to the field of electrophoresis with a brief background concerning electrophoretic separation and characterization. Those who wish to obtain an in-depth understanding of the theory or detailed practical techniques are referred to textbooks/monographs on this subject [1-6]. Next, the advantages of gel electrophoresis as an analytical tool, and the structural requirements regarding dendrimers as electrophoretic analytes are discussed. Finally, studies directed at... 

During electrophoretic separation, the analyte ions displace background co-ions equivalent to their charge. The difference in the conductivity between the analytes and the BGE co-ions induces a signal recorded by the C D detector. It is, therefore, important for optimal sensitivity that the difference in conductance between the analyte and the electrolyte be as high as possible. On the other hand, optimal efficiency of separation is attained when the /tBCE and /tanaiyte are matched. Howevei the ji and the equivalent conductance leqmv are linked ... 

Techniques have also been developed for the specific visualization of particular classes of enzymes following electrophoretic separation in a gel. These techniques are often referred to as activity staining, as the intrinsic activity of the enzyme is used, either to produce a colored product or to produce a clear zone on a colored background within the gel. A method for visualizing proteinases based on the work of Gar-cfa-Carreno and Haard (1993) and Garcia-Car-reno et al. (1993) is presented (see Basic Protocol 3). 

Capillary affinity electrophoresis (CAE) or affinity capillary electrophoresis (ACE) — An electrophoretic separation technique (- electrophoresis), in which -> analytes are separated in a capillary, with the -> supporting (background) electrolyte containing substances capable of specific, often biospecific, interactions with the analytes. Ref [i] Riekkola ML, Jonsson jA, Smith RM (2004) Pure Appl Chem 76 443... 

Fig. 16. Electrophoretic separation of the lipase and esterase activities of porcine pancreas (146, 147). Starch columns equilibrated with 0.025 M acetate buffer, pH 5.25. The activities of the fractions have been determined (o) on emulsions of triolein and tributyrin (black circles), methyl oleate, methyl laurate, and p-nitro-phenyllaurate (black triangles), (b) On solutions of methyl butyrate and p-nitro-phenylacetate (crosses). White circles and dotted line, protein background. Figures along the first peak give the specific activity (lipase) of some fractions, determined against triolein emulsion. Ordinates and abscissas are the same as in Fig. 14.

One of the preferred methods for analyte detection in microfiuidic devices is based on the phenomenon of chemiluminescence (CL), which offers a simple but sensitive means of monitoring low level analyte concentrations [26]. CL reactions typically involve the formation of a metastable reaction intermediate or product in an electronically excited state, which subsequently relaxes to the ground state with the emission of a photon. CL is particularly attractive for portable microfiuidic assays, because the CL reaction acts as an internal light-source, thereby lowering instrumental requirements and significantly reducing power consumption and background interference compared to fluorescence assays. CL-based systems have been successfully applied to on-chip electrophoretic separation of metal ions, immunoassays, and enzyme assays [27], and consequently there is considerable interest in... 

The source and destination vials as well as the inside of the capillary are filled with a buffer, also referred to as carrier electrolyte or background electrolyte. The purpose of the buffer is to maintain the pH as well as the conductivity during the electrophoretic separation. A controlled pH is crucial for maintaining a constant net charge on the biomolecules and, thus, maintaining their electrophoretic mobility A controlled conductivity is required so that Joule heating can be controlled. Buffer concentrations in CE are typically in the order 10-100 mM. 

Williams, R.L., Childs, B., Dose, E.V., Guiochon, G, and Vigh, G, Peak shape distortions in the capillary electrophoretic separations of strong electrolytes when the background electrolyte contains two strong electrolyte co-ions. Anal. Chem., 69,1347,1997. 

Vigh, Gy. Sokolowski, A.D. Capillary electrophoretic separations of enantiomers using cyclodextrin-containing background electrolytes. Electrophoresis 1997,18, 2305-2310. 

MEKC is usually used as a separation technique in which the basic properties of micellar liquid chromatography and CE are combined. MEKC was first described by Terabe in 1984 for the separation of nonionic aromatic compounds and is a powerful separation technique for lipophilic and nonionic species. By addition of surfactants to the background electrolyte, new options for solving electrophoretic separation problems are opened, but it is also possible to apply this technique to study the affinities of drug molecules to surface-active compounds. The term micellar affinity capillary electrophoresis (MACE) is used... 

Busby BM, Vigh G (2005) Synthesis of heptakis(2-0-methyl-3-0-acetyl-6-0-sulfo)-cyclomaltoheptaose, a single-isomer, sulfated P-cyclodextrin carrying nonidentical substituents at all the C2, C3, and C6 positions and its use for the capUlaty electrophoretic separation of enantiomers in acidic aqueous and methanolic background electrolytes. Electrophoresis 26 1978-1987... 

Resolution depends upon differences in mobilities of the species. Background electrolyte of low ionic strength is advantageous, not only to increase electrophoretic (solute) mobilities, but also to achieve low electrical conductivity and thereby to reduce the thermal-convection current for any given field [Finn, in Schoen (ed.), New Chemical Engineering Separation Teehniques, Interscience, New York, 1962]. 

In CZE, the capillary, inlet reservoir, and outlet reservoir are filled with the same electrolyte solution. This solution is variously termed background electrolyte, analysis buffer, or run buffer. In CZE, the sample is injected at the inlet end of the capillary, and components migrate toward the detection point according to their mass-to-charge ratio by the electrophoretic mobility and separations principles outlined in the preceding text. It is the simplest form of CE and the most widely used, particularly for protein separations. CZE is described in Capillary Zone Electrophoresis. ... 

---