Electrophoresis microscopic method

If the individual particles of a colloidal suspension are visible the microscopic method may be used for studying electrophoresis. For this purpose quite a number of arrangements have been utilized. A typical... 

In this section, we shall first give a brief review of the phenomenological theory of these effects.5 -6 26 We shall then show how the methods we have discussed in the previous sections may be extended to derive a microscopic theory of the relaxation effect the microscopic theory of electrophoresis will be considered in the next section. 

Another useful physical property of the crystal is its density, which can be used to determine several useful microscopic properties, including the protein molecular weight, the proteinlwater ratio in the crystal, and the number of protein molecules in each asymmetric unit (defined later). Molecular weights from crystal density are more accurate than those from electrophoresis or most other methods (except mass spectrometry) and are not affected by dissociation or aggregation of protein molecules. The proteinlwater ratio is used to clarify electron-density maps prior to interpretation (Chapter 7). If the unit cell is symmetric (Chapter 4), it can be subdivided into two or more equivalent parts called asymmetric units (the simplest unit cell contains, or in fact is, one asymmetric unit). For interpreting electron-density maps, it is helpful to know the number of protein molecules per asymmetric unit. 

Good descriptions of practical experimental techniques in conventional electrophoresis can be found in Refs. [81,253,259]. For the most part, these techniques are applied to suspensions and emulsions, rather than foams. Even for foams, an indirect way to obtain information about the potential at foam lamella interfaces is by bubble electrophoresis. In bubble microelectrophoresis the dispersed bubbles are viewed under a microscope and their electrophoretic velocity is measured taking the horizontal component of motion, since bubbles rapidly float upwards in the electrophoresis cells [260,261]. A variation on this technique is the spinning cylinder method, in which a bubble is held in a cylindrical cell that is spinning about its long axis (see [262] and p.163 in Ref. [44]). Other electrokinetic techniques, such as the measurement of sedimentation potential [263] have also been used. 

This test, called the Comet Assay or single-cell gel-electrophoresis assay, allows the degree of DNA damage to be determined within a nucleic cell population. The principle of the method is based on the microelectrophoresis of nuclei of isolated cells, under basic conditions, on agarose gel (the whole being observed under a fluorescence microscope). 

Isoelectric focusing and electrophoresis were used for protein mapping and to study protein extraction in horizontal ultra-thin-layer format [138], The 0.12-0.36-mm-thick polyacrylamide gel layer was deposited onto tiny glass plates (e.g., microscope slides). The method enabled the analysis of 1-ng tissue culture specimens. Ultra-thin-layer polyacrylamide isoelectric focusing gel was also employed in two-dimensional analysis of plant and fungal proteins. Marlow et al. [139] reported on the use of 0.2-mm semirigid backing (polyester)-supported... 

A number of methods for the determination of electrophoretic velocity and electrokinetic potential of particles have been developed. These methods include the moving boundary method (a direct study of motion of the boundary between the disperse system and the free dispersion medium due to the applied potential difference), microelectrophoresis (a direct observation of moving particles using a microscope or ultramicroscope), electrophoresis in gels, paper electrophoresis, etc [ 13]. These methods are broadly used to study disperse systems formed with low molecular weight substances, as well as polymers, especially those of natural origin. Electrophoretic methods allow one to separate and analyze mixtures of proteins, and thus are effectively used in scientific research and medical diagnostic applications. 

The lateral mobility of proteins and lipids in natural and artificial lipid bilayer membranes was determined by different methods. For long-range mobility, fluorescence recovery after photobleaching (13-15) and electrophoresis of membrane components (16) were employed. We employed the electrophoresis method for determination of the eletrophoretic and diffusional mobilities of PSI in the plane of hypotonically inflated, spherical thylakoid vesicles. To monitor the redistribution of PSI particles, we made use of the spatial characteristics of the contribution of PSI particles to electrophotoluminescence (EPL) (17, 18). The contribution of PSII to EPL was eliminated by heat treatment of the chloroplasts (19). The EPL originates from the PSI particles at the hemisphere of the vesicles at which the induced electrical field destabilizes the photoinduced charge separation (18). The electrophoretic and diffusional mobilities were measured in vesicular suspensions to avoid immobilization for microscopic visualization (20). The photosynthetic membranes are devoid of cytoskeletal elements that might interfere with the lateral mobility. 

Electrophoresis refers to the motion of a charged particle in a solution in response to an applied electric field. The electrophoresis technique has been widely used to characterize the electrokinetic properties of charged particle-liquid interfaces. In the electrophoresis method, fine particles (usually of 1 pm in diameter) are dispersed in a solution. Under an applied electric field, the particle electrophoresis mobility, vg, defined as the ratio of particle velocity to electric field strength, is measured using an appropriate microscopic technique. The particle -potential is determined from the measured electrophoresis mobility, ve, by using the Smoluchowski equation expressed as... 

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