Electrophoresis heat dissipation

Packed capillaries with a larger inner diameter may be useful in preparative separations. These will find an application in proteome research as a part of multidimensional separation systems that will replace 2-D gel electrophoresis. The preparative CEC will require solving of the problems related to heat dissipation since the radial temperature gradient negatively affects the separations, and sample injection. The fabrication of sintered frits in larger bore capillaries is also very difficult. However, in situ polymerized monolithic frits can be fabricated in capillaries of virtually any diameter [190]. 

Fused silica capillaries are almost universally used in capillary electrophoresis. The inner diameter of fused silica capillaries varies from 20 to 200 pm, and the outer diameter varies from 150 to 360 pm. Selection of the capillary inner diameter is a compromise between resolution, sensitivity, and capacity. Best resolution is achieved by reducing the capillary diameter to maximize heat dissipation. Best sensitivity and sample load capacity are achieved with large internal diameters. A capillary internal diameter of 50 pm is optimal for most applications, but diameters of 75 to 100 pm may be needed for high sensitivity or for micropreparative applications. However, capillary diameters above 75 pm exhibit poor heat dissipation and may require use of low-conductivity buffers and low field strengths to avoid excessive Joule heating. 

Electrophoresis in narrow bore tubes, as performed by Hjerten in 1967, provides a better heat dissipating system. He described an application using glass tubes with an internal diameter (I.D.) of +3 mm. The small volume of the narrow bore tube improves the dissipation of heat due to a lower ratio of the inner volume to the wall surface of a tube (Equation (1)). The better the heat dissipation the higher will be the separation efficiency ... 

A major advantage of capillary gel electrophoresis is that resolution is maintained with increasing field strength, owing to efficient heat dissipation. Introduction of the sample into the gel-filled capillary is typically done electrokinetically (Chapter 6), because hydrostatic injection is not possible with capillaries blocked with gel. Typical injection times between 1 and 20 sec are used at field strengths between 100 and 400 V/cm.28 Coated capillaries should be used to prevent the EOF from pumping the gel out of the capillary. 

The use of exceedingly thin capillary tubes ( 50 /im) in capillary electrophoresis not only reduces convection but it also provides rapid heat dissipation. Using up to 30,000 V, on the order of 106 or more theoretical plates are realized in such tubes (see Figure 8.2). 

Capillary electrophoresis is an instrumental technique for automating electrophoretic analyses.66 It is carried out in fused silica capillaries with internal diameters of 25-100 tm. The capillaries are coated with an external layer of polyimide for mechanical strength. The small internal diameter and high surface-to-volume ratio allow for efficient heat dissipation from the capillaries, enabling separations to be carried out at very high field strengths (up to 1000 V/cm). 

This heat must be dissipated by cooling, which can be done but only to a limited extent The ability to dissipate heat efficiently is usually the factor that limits the speed of electrophoresis, since excess heat leads to non-uniform electrophoresis and a decrease in resolution. The main reason for this is convection in matrix-free electrophoresis in solution, and the effect of temperature on viscosity and diffusion. High temperatures can also lead to denaturation of proteins and nucleic acids. The thinner the layer used for electrophoresis, the more readily is the heat dissipated, and the higher the voltages that can be used. The thickness of the layer will be a compromise between a desire to have a thin layer to minimise heat problems whilst maintaining sufficient capacity to ran samples that can be detected easily. Consis-... 

Fig. 7.7. Modified design of cylindrical gel analytical electrophoresis apparatus. One of the design features, which may also be embodied in the circular layout (Fig. 7.6) is that the tubes are immersed in the lower reservoir buffer for better heat dissipation. A further advantage in this system is the presence of a barrier which impedes the diffusion of electrode products into the buffer solution. 

Slab PAGE allows simultaneous electrophoresis of a number of samples to be performed under identical conditions. Slab PAGE has a higher resolving power than column PAGE, and is often used for molecular weight and purity determinations. Because the slab is relatively thin, heat dissipation is more efficient in a slab than in a column. The slab can be used vertically or horizontally, but in practice, the horizontal slab method is used only when the monomer plus cross-linker concentrations are low and the gel is soft. A slab gel is prepared in the same way as a column gel, except that a comb, or slot-former, is inserted before polymerization. Removal of the comb after the gel has set leaves sample wells that are separated from each other by continuous strips of gel. 

At the other size extreme are electrophoretic "instruments" for separating large quantities (grams to Kilograms) of materials for preparative purposes. Here, due to the large size of the equipment, heat dissipation can become a major problem to be dealt with. More information on electrophoresis on a preparative scale can be found in chapter 16. 

This chapter discusses the phenomena of Joule heating, heat dissipation, estimation of the electrolyte temperature and its effects, and extends the discussion of Nelson and Burgi [3] provided in the first edition of this book. The focus of this chapter is on the use of simple methods to determine the temperature of the electrolyte, free from the influence of Joule heating. An improved understanding of the radial temperature profile that exists during electrophoresis [8] allows a simple technique to be introduced for evaluating the heat transfer coefficient for an instrument. 

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