Magnetic Nanoparticles for Biomedicine - Intracellular Delivery: Fundamentals and Applications - page 369

a clinical-scale level in a closed and sterile system. The key components of the
CliniMACS Plus Instrument are the integrated microcomputer, the magnetic sepa-
ration unit, the peristaltic pump and various pinch valves. The Isolex @ 300 System
(Baxter, USA) is a semi-automated magnetic cell separation system designed to
select and isolate CD34 positive cells, ex vivo, from mobilized peripheral blood
using anti-CD34 monoclonal antibody and superparamagnetic microspheres
(Dynabeads). The device consists of an instrument for separating Dynabeads from
mobilized peripheral blood mononuclear cell (MNC) suspensions and an associated
disposable set for providing the fluid path.
IMS of important pathogenic bacteria ( Salmonella, Legionella, Listeria and
verotoxigenic strains of Escherichia coli (e.g., O157, EPEC/VTEC O103, O111
and O145)) and protozoan parasites ( Cryptosporidium, Giardia ) is efficiently used
in medical, food and water microbiology. Both micrometer and nanometer scale
magnetic particles with immobilized specific antibodies are used as labels. IMS
enables to shorten the detection time which is a very important feature from the
medical point of view. As an alternative to IMS, magnetic particles with immobi-
lized lectins have been used for similar purpose (Šafařík and Šafaříková 1999 ).
Magnetic particles with immobilized annexin V have been employed for the
simple and efficient separation of apoptotic cells from normal culture. This procedure
is based on the fact that annexin V is a Ca 2+ -dependent phospholipid binding protein
with high affinity for phosphatidylserine (PS), which is redistributed from the inner
to the outer plasma membrane leaflet in apoptotic or dead cells. Once on the cell
surface, PS becomes available for binding to annexin V and any of its conjugates.
Magnetic targeting of nucleic acids bound to magnetic particles into the recipient
cells is the basis of magnetofection which is a simple and highly efficient transfec-
tion method. The magnetic iron oxide nanoparticles are usually coated with specific
cationic molecules which can associate with the gene vectors (DNA, siRNA, ODN,
virus, etc.). The magnetic particles are then concentrated on the target cells by the
influence of an external magnetic field generated by magnets. The cellular uptake
of the genetic material is accomplished by endocytosis and pinocytosis, two natural
biological processes. Consequently, membrane architecture and structure stay
intact, in contrast to other physical transfection methods that damage the cell mem-
brane. The nucleic acids are then released into the cytoplasm by different mecha-
nisms depending upon the formulation used. Coupling magnetic nanoparticles to
gene vectors of any kind results in a dramatic increase of the uptake of these vectors
and consequently high transfection efficiency. Transfected cells can be separated
from the non-transfected ones using an appropriate magnetic separation technique
(Plank et al. 2003 ; Scherer et al. 2002 ; Safarik and Safarikova 2009a ).
Magnetic separation of nucleic acids and proteins enables direct isolation and
purification of target biomolecules from difficult-to-handle biological samples,
such as cell homogenates or body fluids. Magnetic separation techniques have
several advantages in comparison to traditional separation procedures; the laboratory-
scale process is very simple, and all steps of the purification can take place in one
test tube without expensive liquid chromatography systems (Berensmeier 2006 ;
Šafařík and Šafaříková 2004 ).