Continuous-flow electrophoresis discussion

In the second step of intrinsic thromboplastin formation, active PTA reacts with inactive plasma thromboplastin component (PTC) to convert it to active PTC. The active PTC reacts with AHG and factor X to yield a compound referred to here as product I. What is substrate and what is enzyme in this reaction are not known. A little is known of product I because it has been possible to purify it chromatographically on DEAE and on continuous flow electrophoresis. Its properties will be discussed below. 

The theory of electrophoresis has been adequately covered in the excellent textbooks of Giddings [1] and Andrews [2] as well as in specific manuals [3], [4]. For discussion on electrophoresis in free liquid media, e.g., curtain, freeflow, endless belt, field-flow-fractionation, particle, and cell electrophoresis the reader is referred to a comprehensive review by Van Oss [5] and to a book largely devoted to continuous-flow electrophoresis [6], Here the focus is mostly on electrophoresis in a capillary support, i.e, in gel-stabilized media, and discussion is limited to protein applications. 

The author would like to acknowledge R S Technologies, Inc. (Wakefield, RI, USA) for the loan of the continuous free flow electrophoresis system, and Cerestar, Inc. for the donation of the sulfated cyclodextrin. The author would also like to thank Drs. Chris Welch and Prabha Painuly for helpful discussions. 

It is apparent from the above sections that the understanding of electrophoretic mobility involves both the phenomena of fluid flow as discussed in Chapter 4 and the double-layer potential as discussed in Chapter 11. In both places we see that theoretical results are dependent on the geometry chosen to describe the boundary conditions of the system under consideration. This continues to be true in discussing electrophoresis, for which these two topics are combined. As was the case in Chapters 4 and 11, solutions to the various differential equations that arise are possible only for rather simple geometries, of which the sphere is preeminent. 

Mass spectrometry is a valuble tool with which an abundancy of structural information may be obtained from a minute amount of material. Capillary electrophoresis may be interfaced with mass spectrometry by electrospray ionization [124-126] or continuous-flow, fast-atom bombardment methods [127,128]. Several reviews discuss applications of the interfacing techniques, and address the attributes and disadvantages associated with these methods [129,130]. Critical parameters involved in the optimization of CE-electrospray ionization mass spectrometry analysis have been reviewed as well [131],... 

Li and Harrison carried out the first cell assay in microchannels [2]. This seminal work made use of electrokinetically driven flow (electroosmosis and electrophoresis) to transport bacteria, yeast, and mammalian cells in channels and to implement low-volume chemical lysis (cell death). This theme of microfluidics-based cell transport, sorting, and lysis, has continued to be a popular application, as well as related work in using microfluidics to culture cells and to pattern them into structures. We acknowledge the utility of these methods (and note that they are featured in several good reviews - see El-Ali et al. [1] and other entries in the Encyclopedia), but focus here on describing microfluidics-based cell assays that fit the definition described above - application of a stimulus and measurement of a response. These assays fall into four broad themes sorted as a function of the t)fpe of response to be measured intracellular biochemistry, extracellular biochemistry, mechanical properties, and electrical properties. Prior to discussing these kinds of assays, we describe the basic methodology common to all forms of cell assays in microfluidics devices. 

This last point needs some elaboration. Consider in Figure 7.1.29(c), any one of tbe two ports used to withdraw two different protein products continuously. If we bave three proteins present in the feed stream, then let the liquid coming to one of the ports be pure in one protein it is clear that the liquid passing over the other port will have two other proteins therefore the liquid stream withdrawn from this port cannot be pure in one protein. This is an inherent limitation of processes where the bulk flow is parallel to the direction of the force. Here, even though we have one force perpendicular to the bulk flow, in the presence of the electrical force parallel to bulk flow, the system in continuous operation is just like the counter-current electrophoresis of Figure 6.3.4 and equations (6.3.9a,b). However, in a hatch mode, one can have multi-component separation since each species will be focused to its own zone and therefore can be withdrawn at a later time. Multicomponent separation in electrochromatography needs a different flow vs. force configuration, to he discussed further in Section 8.2. 

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