Fractionation migration

The proteins known today as linker or HI histones were initially described as the abundant lysine-rich nuclear proteins that could be separated by chromatography on ion exchange resin from other major basic nuclear proteins known today as core histones (for review see Refs. [1,2]). During gel electrophoresis of histones the HI fraction migrated as the slowest and most heterogeneous band. Upon the discovery of nucleosomal organization of chromatin in the mid 1970s it turned out that linker histones are not involved in the assembly of the nucleosomal protein core, but bind to DNA between nucleosomes (hence their name). 

An important advantage of two-dimensional methods is the solitary migration of each fraction on its own path. In contrast with unidimensional or zone electrophoresis where each fraction migrates its own distance but over a common path, no contamination of fractions is possible (Fig. 40). If the substrate is sufficiently long and wide and a high intensity electrical field is applied, it becomes possible to separate fractions reasonably completely from each other. If the physical forces involved are deployed skillfully, the electrical field can be driven up so that rapid and wide separations are obtained. This point is especially important in star electrophoresis, whereas in collecting electrophoresis mastery of the physical forces provides an excellent continuous preparative tool. 

Now calculating the fraction migrated from the polymer into the liquid ... 

Then calculating the fraction migrated using the mass balance equation ... 

The fraction of migrant diffused from F into P up to time t, from mpo = VpCpo and the fraction migrated from P into F up to time t, from mpo = FpCpo are ... 

Fig-1 Fractional migration X L) of solute bands along the column and resulting chromatograms for isocratic versus gradient elution. (Reprinted with permission from L. R. Snyder, M. A. Stadalius, and M. A. Quarry, Analytical Chemistry,... 

Flo. 42. Fractional migration of proteins during gradient elution. Colored proteins separated in glass-walled colunte From Di Bussolo and Gant (779). 

Pig pancreatic juice lipase has been purified by a similar method. The degree of purification and properties of the purified lipase from the juice are similar to those of the preparation from pancreatin (223). Borgstrom (225) subjected rat pancreatic juice to electrophoresis and found that the lipase activity was concentrated in a small fraction migrating toward the cathode. Purification of about 40-50 times was obtained. 

This can be viewed as a dynamic equilibrium in which the anion spends a certain time fraction migrating as the free anion and another time fraction as the association complex, during which migration is much slower. The magnitude of this effect is a function of the type and concentration of the electrolyte cation, the counter-anion and of the properties of the analyte anion. 

Figure 11.27 Mass fraction migrated from a polymer film or sheet.

Because of the lower rotation speeds in a disc centrifuge a different analysis technique has to be employed. The cell is first filled with a spin fluid and then a sample layer is injected on top of the fluid while the disc is rotating. By this means the particle fractions migrate past the detection optics layer by layer according to their differing sedimentation velocities. Unfortunately, it is often difficult to achieve a uniform injection layer in practice (because of disruptions of the sample flow front). For this reason, and also because of the low density difference between the polymer particles and the aqueous phase, disc centrifuge sedimentometry is not widely used for the characterization of polymer dispersions. 

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