The simplest technique for adding ferromagnetic properties to biological
matter is to supplement the solution with small magnetite particles and
using the natural surface and colloidal properties of both the magnetite and
the biological material to obtain the desired separation from the solution
[1]. Although this technique is not very selective, it works very well in cases
where total removal of cell mass is required, for example, from fermentation
broth or waste water. The technique can be further improved by adjusting
surface and/or colloidal properties and/or by adding a flocculating agent [6].
Ferrofluids (surfactant stabilized magnetite colloids) had also been extensively
used for ferromagnetic labeling as reviewed by Rosenberg [20] and their
application in affinity chromatography was discussed by Mosbach and
Anderson [21].
Most of the applications of HGMS of paramagnetic material pertain to red
blood cells (erythrocytes) [6, 22]. The technique relies on the paramagnetic
properties of hemoglobin groups in erythrocytes when they are in the reduced
state (Fe 2+ ). When hemoglobin combines with oxygen it becomes diamagnetic.
No evidence of damage to erythrocytes was found when they were separated
from whole blood by HGMS [22]. This opened up a wide range of clinical and
research uses of HGMS of red blood cells as this method has advantages over
centrifugation in that it is completely selective and can be made a continuous
process. The low magnetic susceptibility of erythrocytes [22] is compensated by
using lower flow rates than in the other (magnetically labeled cells) applications
of HGMS to maintain the level of adsorption [1]. There are other ways to induce
paramagnetism. For instance, large positive susceptibilities can be introduced
into biological particles by their adsorption of magnetic cations. The trivalent
cations of the lanthanide series are especially effective at relatively low
magnetizing fields compared to hemoglobin. It was demonstrated that cells
from whole blood, yeast cells, bacteria and Vistna virus can be magnetically
recovered from Er 3+ -containing solutions with an efficiency greater than 80%
[23]. The binding of the ion is affected by the pKa of the binding group, the pH,
and the ionic strength and hence these parameters could be varied to magnetize
a specific material preferentially [1].
Diamagnetic materials can be separated by using a strongly paramagnetic
background fluid which increases the contrast. White blood cells and platelets
have been extracted by this method though in general the range of concentration of the carrier medium is limited by the osmotic pressure effects on the
cell membrane [6].
Two possible modes – direct and indirect – are available for the isolation of
target cells or compounds. The former is often employed, in which magnetic
particles with immobilized affinity ligands are directly suspended in the
solution or suspension. During the course of incubation the target cells bind
with the immobilized ligand and the whole complex can then be separated
using a magnet. In the indirect mode, the target cells interact with the affinity
ligand, usually a primary antibody, that has first been added to the solution or
suspension in the free form, to form a complex during incubation. Magnetic
particles with immobilized secondary antibodies are then added. Finally the
magnetic complex formed is separated in a magnetic separator.
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K.S.M.S. Raghavarao et al.
matter is to supplement the solution with small magnetite particles and
using the natural surface and colloidal properties of both the magnetite and
the biological material to obtain the desired separation from the solution
[1]. Although this technique is not very selective, it works very well in cases
where total removal of cell mass is required, for example, from fermentation
broth or waste water. The technique can be further improved by adjusting
surface and/or colloidal properties and/or by adding a flocculating agent [6].
Ferrofluids (surfactant stabilized magnetite colloids) had also been extensively
used for ferromagnetic labeling as reviewed by Rosenberg [20] and their
application in affinity chromatography was discussed by Mosbach and
Anderson [21].
Most of the applications of HGMS of paramagnetic material pertain to red
blood cells (erythrocytes) [6, 22]. The technique relies on the paramagnetic
properties of hemoglobin groups in erythrocytes when they are in the reduced
state (Fe 2+ ). When hemoglobin combines with oxygen it becomes diamagnetic.
No evidence of damage to erythrocytes was found when they were separated
from whole blood by HGMS [22]. This opened up a wide range of clinical and
research uses of HGMS of red blood cells as this method has advantages over
centrifugation in that it is completely selective and can be made a continuous
process. The low magnetic susceptibility of erythrocytes [22] is compensated by
using lower flow rates than in the other (magnetically labeled cells) applications
of HGMS to maintain the level of adsorption [1]. There are other ways to induce
paramagnetism. For instance, large positive susceptibilities can be introduced
into biological particles by their adsorption of magnetic cations. The trivalent
cations of the lanthanide series are especially effective at relatively low
magnetizing fields compared to hemoglobin. It was demonstrated that cells
from whole blood, yeast cells, bacteria and Vistna virus can be magnetically
recovered from Er 3+ -containing solutions with an efficiency greater than 80%
[23]. The binding of the ion is affected by the pKa of the binding group, the pH,
and the ionic strength and hence these parameters could be varied to magnetize
a specific material preferentially [1].
Diamagnetic materials can be separated by using a strongly paramagnetic
background fluid which increases the contrast. White blood cells and platelets
have been extracted by this method though in general the range of concentration of the carrier medium is limited by the osmotic pressure effects on the
cell membrane [6].
Two possible modes – direct and indirect – are available for the isolation of
target cells or compounds. The former is often employed, in which magnetic
particles with immobilized affinity ligands are directly suspended in the
solution or suspension. During the course of incubation the target cells bind
with the immobilized ligand and the whole complex can then be separated
using a magnet. In the indirect mode, the target cells interact with the affinity
ligand, usually a primary antibody, that has first been added to the solution or
suspension in the free form, to form a complex during incubation. Magnetic
particles with immobilized secondary antibodies are then added. Finally the
magnetic complex formed is separated in a magnetic separator.
146
K.S.M.S. Raghavarao et al.