2.2
Electrophoretic Extraction
2.2.1
Existing Methods – Brief Analysis
Electrophoresis is a leading method for resolving mixtures of charged macromolecules (either proteins or nucleic acids) or cells. The electrophoretic separation of proteins without gels has been a long-standing goal of separation
research [45, 46]. Electrophoretic separations are influenced by many factors
including the size (or molecular weight), shape, secondary structure, and
charge of the macromolecule or cell. These features can influence electrophoretic properties either separately or jointly. In order to achieve the required
scale-up scientists and engineers resort to flowing methods [46–48]. Scale-up
of electrophoresis is hindered by ohmic heating. The heat generated is equal to
the product of the current and voltage, and this heat can cause free convection
and mixing within the system. Too much heat can also denature the labile biomolecules or cells.
Decades of research in free electrophoresis have identified thermal convection [49], electro-osmosis [50], particle sedimentation [51], droplet sedimentation [52], particle aggregation [53], and electro-hydrodynamic zone
distortion [54] as the major obstacles to scale-up. Free electrophoresis has not
gained popularity as a preparative or industrial separation method owing to
these gravity-dependent characteristics. Density gradients [52, 55] or elaborate
flowing devices have been required up to now to stabilize free fluid systems
and/or the particles suspended in them while they are non-isothermally heated
by the passage of an electric current. Without the need to prepare density
gradients and/or use elaborate flowing systems, free electrophoresis could enjoy
more widespread use, because it is a high-resolution separation method that
does not require adsorption to solid media and the subsequent solids handling.
Furthermore, it can handle particles (cells) as well as solutes (macromolecules)
alike. To name a few, specific applications of free electrophoresis include the
separation of different cells of peripheral blood and bone marrow in
hematological and immunological research and potentially in clinical
therapeutic applications [56], and the separation of proteins from body fluids,
tissue extracts, and fermentation broths in biotechnology [57].
None of these principal gravity-dependent and gravity-independent processes have been investigated in multistage electrophoresis, which is designed to
minimize the impact of these processes on separation quality. All of these
processes have been studied in the past in applications to other forms of free
electrophoresis as reviewed by Todd [34, 41].
Thermal convection, which in turn causes mixing, is induced by ohmic
heating when current passes through the buffer and heats the buffer non-uniformly [49]. Above a critical Rayleigh number (Ra c ), convective circulation sets
in, and it occurs in both static and flowing electrophoresis systems [54, 58]. The
Ra c is unexpectedly low in certain flowing electrophoresis applications, so that
thermal convection is a significant deterrent to the development of electroMultistage Magnetic and Electrophoretic Extraction of Cells, Particles and Macromolecules
157
Electrophoretic Extraction
2.2.1
Existing Methods – Brief Analysis
Electrophoresis is a leading method for resolving mixtures of charged macromolecules (either proteins or nucleic acids) or cells. The electrophoretic separation of proteins without gels has been a long-standing goal of separation
research [45, 46]. Electrophoretic separations are influenced by many factors
including the size (or molecular weight), shape, secondary structure, and
charge of the macromolecule or cell. These features can influence electrophoretic properties either separately or jointly. In order to achieve the required
scale-up scientists and engineers resort to flowing methods [46–48]. Scale-up
of electrophoresis is hindered by ohmic heating. The heat generated is equal to
the product of the current and voltage, and this heat can cause free convection
and mixing within the system. Too much heat can also denature the labile biomolecules or cells.
Decades of research in free electrophoresis have identified thermal convection [49], electro-osmosis [50], particle sedimentation [51], droplet sedimentation [52], particle aggregation [53], and electro-hydrodynamic zone
distortion [54] as the major obstacles to scale-up. Free electrophoresis has not
gained popularity as a preparative or industrial separation method owing to
these gravity-dependent characteristics. Density gradients [52, 55] or elaborate
flowing devices have been required up to now to stabilize free fluid systems
and/or the particles suspended in them while they are non-isothermally heated
by the passage of an electric current. Without the need to prepare density
gradients and/or use elaborate flowing systems, free electrophoresis could enjoy
more widespread use, because it is a high-resolution separation method that
does not require adsorption to solid media and the subsequent solids handling.
Furthermore, it can handle particles (cells) as well as solutes (macromolecules)
alike. To name a few, specific applications of free electrophoresis include the
separation of different cells of peripheral blood and bone marrow in
hematological and immunological research and potentially in clinical
therapeutic applications [56], and the separation of proteins from body fluids,
tissue extracts, and fermentation broths in biotechnology [57].
None of these principal gravity-dependent and gravity-independent processes have been investigated in multistage electrophoresis, which is designed to
minimize the impact of these processes on separation quality. All of these
processes have been studied in the past in applications to other forms of free
electrophoresis as reviewed by Todd [34, 41].
Thermal convection, which in turn causes mixing, is induced by ohmic
heating when current passes through the buffer and heats the buffer non-uniformly [49]. Above a critical Rayleigh number (Ra c ), convective circulation sets
in, and it occurs in both static and flowing electrophoresis systems [54, 58]. The
Ra c is unexpectedly low in certain flowing electrophoresis applications, so that
thermal convection is a significant deterrent to the development of electroMultistage Magnetic and Electrophoretic Extraction of Cells, Particles and Macromolecules
157