This document summarizes research on using magnetic nanoparticles functionalized with antibodies to target and separate tumor cells from other cells. The nanoparticles were conjugated with anti-MICA antibodies, which target a ligand on HeLa cells. HeLa cells were infected with the nanoparticles and detached using trypsin-EDTA. A magnetic field was then used to separate the HeLa cells from other cells due to the magnetic nanoparticles attached to their surface via the antibodies. Transmission electron microscopy confirmed the nanoparticles had been endocytosed by the HeLa cells through receptor-mediated endocytosis. This targeted magnetic separation technique can isolate tumor cells from cell cultures or samples.

1. Specific magnetic cell separation using
Grupo de Investigación
Universidad de Oviedo receptor-mediated endocyted iron oxide y Desarrollo de
Materiales Magnéticos
1 1 1 2 3 3 4 1
D. Lago-Cachón , L. Elbaile , M. Rivas , A.M. López-Quintela , C. López-Larrea , A. López-Vázquez , G. Martínez-Paredes , J.A. García
1
University of Oviedo, Dept. of Physics, CCalvo Sotelo s/n, 33007 Oviedo, Spain.
2
University of Santiago de Compostela, Dept. of Physical Chemistry, E-15782 Santiago de Compostela, Spain.
3
Histocompatibility and Transplantation Unit, Hospital Universitario Central de Asturias, 33006 Oviedo.
4
Healthsens S.L., CEmilio rodríguez Vigil s/n—Bl Polivalente A, 4ª planta, 33006 Oviedo, Spain.
Introduction
The aim of the project, in which this work is
included, is in vitro labelling, magnetic separation
and detection of tumor cells.
Functionalized magnetite nanoparticles are
bound to specific antibody against a surface ligand
of the tumor.
Finally, the nanoparticles could be detected by a
magnetic biosensor based on the Giant Magneto
Impedance effect of an amorphous metallic ribbon.
Fabrication and functionalization of nanoparticles
10 nm mean size magnetite nanoparticles dispersed in water
were used. They were produced in Nanogap using a co-
precipitation method (NGAP NP FeO 2204-W).
The nanoparticles surface was modified with EDC and sulfo-NHS
in order to provide free amine groups where streptavidin could be
attached. Finally, biotinylated antibodies were added. Anti-human
MICA was used, because of its strong relation with antitumor
immunity.
HeLa cells growing on surface. Nanoparticles are not here visible,
except for the agglomerate indicated with a circle.
Avoiding agglomeration of nanoparticles is of great importance.
When functionalization is properly done, nanoparticles’
hydrodynamic diameter is smaller than 200 nm, and are not visible
by optical microscopy. When agglomeration occurs, magnetite
particles are visible and could not be endocyted.
Cell culture and infection
2
HeLa cells were cultured in DMEM over 25 cm flask surface
until 75% of confluence were achieved.
Cells were washed and infected with 0.1 mg of functionalized
nanoparticles. After 30’ conjugation time, cells were detached in
trypsin-EDTA and magnetically separated.
HeLa cells in suspension after conjugation. Nanoparticles are not
visible. Roughness of cells surface is not related to them.
Video of magnetic cell separation, using a modified
microscope to allow application of magnetic field gradient,
and study of cells kinematics.
URL: https://plus.google.com/photos/113013190367394546907/albums/5712259340294179265?authkey=CKS7uemThc-EwAE
Different cells lines with no MICA expression.
Separated HeLa cells were analyzed by transmission electron
microscopy, showing that the nanoparticles were receptor-
mediated endocyted by HeLa cells.
Conclusion
Results show that magnetic nanoparticles tag specifically HeLa cells
which can be thus concentrated by means of a magnetic field gradient.
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