1
Soft magnetic materials: from microsensors to cancer therapy
Alfredo García-Arribas
NEW MATERIALS FOR SENSORS AND ACTUATORS
DATE: September 28, 2017
TIME: 9:30h – 13:00h
VENUE: Conference Room – Jeronimo de Ayanz Building – C/Tajonar 22 (Public University of Navarra)
ABSTRACT
Scientific and technological breakthroughs in new advanced materials are revolutionizing many industrial and consumer products and are the platforms for continued innovation in many rapidly growing industrial sectors. In particular, many devices are based on new materials properties and processing techniques. These sensors and actuators are being used in environment control, bioelectronics, nanoelectronics, agricultural strategies, automotive industries etc. Besides, these devices should also fit the requirements necessaries to work in several particular conditions. Among others, shape memory alloys, graphene, hybrid and magnetic materials are promising candidates for actual and future applications.
This workshop is held under the financing agreement signed between the Public University of Navarra and the Obra Social La Caixa – CAN Foundation
PROGRAMME
9:30h Opening Iñaki Pérez de Landazabal – Head of INAMAT Ramón Gonzalo – UPNA Research Vice-rector
9:45h Integrative approaches to Inorganic and hybrid Nanomaterials Clement Sanchez. Laboratoire Chimie de la Matière Condensée de Paris UMR ,CNRS, France
10:30 Chemistry of Novel 2D Materials Beyond Graphene Gonzalo Abellán. Univ Erlangen Nurnberg, Department of Chemistry and Pharmacy, Erlangen, Germany
11:15 11:30- Coffee break 11:30 3D printing, a disruptive technology, challenging creativity
Jan Van Humbeeck. Department of Mechanical Engineering (MECH), KU Leuven, Belgium
12:15h Soft magnetic materials: from microsensors to cancer therapy Alfredo Garcia Arribas. Electricity and Electronics Department. Basque Country University (UPV/EHU) and BC Materials, Leioa, Spain .
13:00 Closure Lunch & networking
Registration needed: https://goo.gl/forms/tPQ2rxIDbGOrSYOY2
New materials for sensors and actuators. Pamplona, 28 septiembre 2017
2
Outline• Introduction
• Magnetism basics
• Magnetization process
• Soft magnetic materials
• Magnetic microsensors • Magnetic field sensors
• Magneto-impedance sensors
• Electronic compass
• Magnetoelastic sensors • Magnetoelastic effect
• Magnetoelastic resonance
• Oil viscosity sensor
• Magnetic nanodiscs for cancer therapy • Magneto-mechanical actuation
• Magnetic vortex state
• Sub-100 nm vortex discs
• Nanodiscs in cancer cells
3
Outline• Introduction
• Magnetism basics
• Magnetization process
• Soft magnetic materials
• Magnetic microsensors • Magnetic field sensors
• Electronic compass
• Magneto-impedance sensors
• Magnetoelastic sensors • Magnetoelastic effect
• Magnetoelastic resonance
• Oil viscosity sensor
• Magnetic nanodiscs for cancer therapy • Magneto-mechanical actuation
• Magnetic vortex state
• Sub-100 nm vortex discs
• Nanodiscs in cancer cells
Magnetism Basics
Atomic magnetic moments
Exchange interaction
Spins sum according to certain rules. Only some atoms display net atomic moment.
Magnetism in matter is a quantum effect. Some ingredients are necessary:
Spin of the electronsIt can have only two orientations: up and down.
Only in few materials these atomic moments aligns themselves spontaneously
ferromagnetic antiferromagnetic ferrimagnetic
For applications, only ferromagnetism is useful. Ferrimagnetism is similar.
4
5
Magnetostatic energyA magnetized material produced a strong
external magnetic field.
Huge magentostatic energy.
Magnetism BasicsCrystal anisotropyExchange is isotropic.
Magnetic moments align in selected directions
according to crystal symmetry.
Magnetic domainsThe magnetization is distributed in domains with
different orientations, compatible with anisotropy
to minimize the magnetostatic energy.
The material is macroscopically no-magnetized
6
Magnetism BasicsDomain wallsThe boundary between domains are called domain walls
Domain walls increase the exchange energy.
The equilibrium configuration is an energy compromise.
Magnetic force microscopy
Kerr effect microscopy
7
Magnetization processWhen a magnetic field is applied, the material is magnetized.
Domain wall movement Rotation of the magentization
M is the magnetic moment per unit volume, measured in the direction of the applied field
Hex
M
8
Hysteresis loop
Domain walls are pinned in defects, grain boundaries, etc.
Hex
M
Hc
Mr
HARD magnetic material SOFT magnetic material
9
Magnetic materialsHard magnetic materialsMagnets.
large Mr: produce external magnetic field.
large Hc: are difficult to demagnetise
Soft magnetic materialsLarge permeability.
Low energy loss (slim loop)
µr =dM
dH
Have large response to small magnetic fields.Concentrate and guide the magnetic flux.
Transformers Electric motors
r
10
Soft magnetic materials
Intrinsic propertiesCrystal structure
Magnetic anisotropyTreatmentsCold working
Thermal annealing
….
The softness of a magnetic material is determined by
Grain-oriented silicon steel for transformers
Permalloy Fe20Ni80 has no crystal anisotropy
The softness can be induced only in the direction of interest
-1.0
-0.5
0.0
0.5
1.0
-10 -5 0 5 10
easyhard
Ker
r sig
nal
H (Oe)
HH
easy axis
Permalloy deposited under applied field
11
Soft magnetic materials
crystalline alloy amorphous alloy
Amorphous magnetic alloys (metallic glasses)Topological (and chemical) disorder.
No pinning for domains walls.
No magnetic anisotropy
Extremely soft!
Rapid quenching preparation methodcooling rates of 106 degrees per second Prepared in the form of ribbon or wires
Fe, Co, Ni and B, P, C, Nb, Zr, etc
12
Outline• Introduction
• Magnetism basics
• Magnetization process
• Soft magnetic materials
• Magnetic microsensors • Magnetic field sensors
• Electronic compass
• Magneto-impedance sensors
• Magnetoelastic sensors • Magnetoelastic effect
• Magnetoelastic resonance
• Oil viscosity sensor
• Magnetic nanodiscs for cancer therapy • Magneto-mechanical actuation
• Magnetic vortex state
• Sub-100 nm vortex discs
• Nanodiscs in cancer cells
13
Magnetic field sensorsThe great sensitivity to small fields makes soft magnetic materials excellent for magnetic field sensing.
Electronic compassMeasure the Earth magnetic field (about 0.5 G or 50 μT).
Together with accelerometers and gyroscopes provide attitude detection.
Magneto-inductive detection
High resolution.
Poor microelectronic integration.
3-Ax
is D
igita
l Com
pass
IC
HMC5
843
The
Hone
ywel
l HM
C584
3 is
a s
urfa
ce m
ount
mul
ti-ch
ip m
odul
e de
sign
ed
for
low
field
mag
netic
sen
sing
with
a d
igita
l int
erfa
ce fo
r app
licat
ions
suc
h
as lo
w co
st c
ompa
ssin
g an
d m
agne
tom
etry
. Th
e H
MC5
843
incl
udes
our
stat
e of
the
art
1043
ser
ies
mag
neto-
resis
tive
sens
ors
plus
Ho
neyw
ell
deve
lope
d AS
IC c
onta
inin
g am
plifi
catio
n, s
trap
driv
ers,
offs
et c
ance
llatio
n,
12-b
it AD
C an
d an
I2 C
ser
ial b
us in
terfa
ce. T
he H
MC5
843
is in
a 4
.0 b
y 4.
0
by 1
.3m
m s
urfa
ce m
ount
lead
less
chi
p ca
rrier
(LCC
). Ap
plica
tions
for
the
HMC5
843
inclu
de
Cons
umer
El
ectro
nics
, Au
to
Navig
atio
n Sy
stem
s,
Pers
onal
Nav
igat
ion
Devic
es, a
nd M
agne
tom
eter
s.
Th
e HM
C584
3 ut
ilizes
Hon
eywe
ll’s A
niso
tropi
c M
agne
tore
sistiv
e (A
MR)
tech
nolo
gy th
at p
rovi
des
adva
ntag
es o
ver o
ther
mag
netic
sen
sor t
echn
olog
ies.
The
sen
sors
feat
ure
prec
ision
in-a
xis
sens
itivi
ty a
nd li
near
ity, s
olid
-sta
te c
onst
ruct
ion
with
very
low
cros
s-ax
is s
ensi
tivity
des
igne
d to
mea
sure
bot
h di
rect
ion
and
mag
nitu
de o
f Ear
th’s
mag
netic
fiel
ds, f
rom
tens
of
micr
o-ga
uss
to 6
gau
ss. H
oney
well’s
Mag
netic
Sen
sors
are
am
ong
the
mos
t sen
sitive
and
relia
ble
low-
field
sen
sors
in th
e
indu
stry
.
Hone
ywel
l co
ntin
ues
to m
aint
ain
prod
uct
exce
llenc
e an
d pe
rform
ance
by
intro
ducin
g in
nova
tive
solid
-sta
te m
agne
tic
sens
or s
olut
ions
. Th
ese
are
high
ly re
liabl
e, t
op p
erfo
rman
ce p
rodu
cts
that
are
del
ivere
d wh
en p
rom
ised.
Hon
eywe
ll’s
mag
netic
sen
sor s
olut
ions
pro
vide
real
sol
utio
ns y
ou c
an c
ount
on.
FEA
TURE
S
BEN
EFIT
S
43-
Axis
Mag
neto
resis
tive
Sens
ors
and
ASIC
in a
Sin
gle
Pack
age
4Sm
all S
ize fo
r Hi
ghly
Inte
grat
ed P
rodu
cts.
Jus
t Add
a M
icro-
Cont
rolle
r Int
erfa
ce, P
lus
Two
Exte
rnal
SM
T Ca
pacit
ors
4Lo
w Co
st
4De
signe
d fo
r Hig
h Vo
lum
e, C
ost S
ensit
ive O
EM D
esig
ns
44.
0 x
4.0
x 1.
3mm
Low
Hei
ght P
rofil
e
LCC
Surfa
ce M
ount
Pac
kage
4Ea
sy to
Ass
embl
e &
Com
patib
le w
ith H
igh
Spee
d SM
T As
sem
bly
4Lo
w Vo
ltage
Ope
ratio
ns (2
.5 to
3.3
V)
4Co
mpa
tible
for B
atte
ry P
ower
ed A
pplic
atio
ns
4Bu
ilt-In
Stra
p Dr
ive C
ircui
ts
4Se
t/Res
et a
nd O
ffset
Stra
p Dr
ivers
for D
egau
ssin
g, S
elf T
est,
and
Offs
et C
ompe
nsat
ion
4I2 C
Dig
ital I
nter
face
4Po
pula
r Two
-Wire
Ser
ial D
ata
Inte
rface
for C
onsu
mer
Ele
ctro
nics
4Le
ad F
ree
Pack
age
Cons
truct
ion
4Co
mpl
ies
with
Cur
rent
Env
ironm
enta
l Sta
ndar
ds
4W
ide
Mag
netic
Fie
ld R
ange
(+/-6
Oe)
4Se
nsor
s Ca
n Be
Use
d in
Stro
ng M
agne
tic F
ield
Env
ironm
ents
4Av
aila
ble
in T
ape
& Re
el P
acka
ging
4Hi
gh V
olum
e O
EM A
ssem
bly
Micro flux-gate Anisotropic magnetoresistance (AMR)
Difficult integration.
Noise problems.
Used in some systems
Need of reset field.
Technologies using soft magnetic materials
Both use the change in permeability Use magnetorresistance
14
Magnetic field sensorsHall sensorsWinner technology.
Use Si Hall effect.
Completely integrable (sensor and logic in the same die).
Low sensibility. Need a magnetic concentrator.
The Permalloy concentrator amplifies the magnetic field and guides the perpendicular components into the planar sensors.
15
Magneto-impedance sensors
Magneto-impedance effectSearch for totally integrable solutions with higher sensitivity.
Hiac(!) r
j (A/m2)
δ
δ: penetration depth
Skin effect: the alternating current flows near the surface of the conductor
� =
r2
!�µ
Zmin
ΔZMI (%) = x 100
H
Z
μhigh
μlowZmin
Z
ΔZ
max
16
Z
H
ΔZ
μhigh
μlow
Zmax
Zmin
max. sensibility
Transverse anisotropyH
M
H
Magneto-impedance sensorsMI in planar samples (i.e. thin films)
• Very large sensitivity at low fields in samples with in-plane, transverse anisotropy.
• Need for thick samples to enhance the skin effect
HH
-1.0
-0.5
0.0
0.5
1.0
-10 -5 0 5 10
easyhard
Ker
r sig
nal
H (Oe)
20 nm thick
-1.0
-0.5
0.0
0.5
1.0
-100 -50 0 50 100
easyhard
Ker
r sig
nal
H (Oe)
260 nm thick
well defined in-plane anisotropy development of out-of-plane anisotropy
Problem: softness lost in thicker samples
17
Magneto-impedance sensors
thin (6 nm) Ti spacers
Py layers below critical thickness
~1 μm
-1.0
-0.5
0.0
0.5
1.0
-10 -5 0 5 10
M/M
s
H (Oe)
0
5
10
15
20
-50 -25 0 25 50
MI (
%)
H (Oe)
f = 200 MHz
MI = 23 %
First strategy: increase thickness using non-magnetic spacers
Magnetic softness preserved MI ratio enhancement
A. Svalov et al. APL 100, 162410 (2012)
18
Magneto-impedance sensorsSecond strategy: enhance MI using the magneto inductive effect
Py
Cu
Py/Ti
Py/Ti
MagneticConductive non-magneticMagnetic
Search for best layer configuration:
• thickness of the layers
• thickness ratio of magnetic to non-
magnetic layers
50
100
150
200
250
300
350
1
2
3
4
ΔZ/
Z (%
)
Z ( Ω)
f = 23 MHz
-300
-150
0
150
300
-2
0
2
-30 -20 -10 0 10 20 30
s (%
/Oe)
dZ/dH (Ω
/Oe)
H (Oe)smax = 300%/Oe (2.7 Ω/Oe = 27 kΩ/T)
Extraordinary performance!
[Py(100 nm)/Ti(6 nm)]4 / Cu(400 nm) / [Ti(6 nm)/Py(100 nm)]4
Best performing multilayer structure is
19
• deposited onto Si substrate
• shaped by photolithography
1
20#!m40#!m60#!m80#!m100#!m120#!m140#!m
0.5#mm 1.0#mm 1.5#mm 2.0#mm
width
length
microstrip coplanar
Electronic compass
Micro-sensors
Magneto-impedance sensors
14.5
15.0
15.5
16.0
16.5
17.0
17.5
0 180 360 540 720 900 1080
Z
( Ω)
θ (º)
θSample
Earth field
s = 25 mΩ/º
20
Outline• Introduction
• Magnetism basics
• Magnetization process
• Soft magnetic materials
• Magnetic microsensors • Magnetic field sensors
• Electronic compass
• Magneto-impedance sensors
• Magnetoelastic sensors • Magnetoelastic effect
• Magnetoelastic resonance
• Oil viscosity sensor
• Magnetic nanodiscs for cancer therapy • Magneto-mechanical actuation
• Magnetic vortex state
• Sub-100 nm vortex discs
• Nanodiscs in cancer cells
21
Magnetoelastic sensors
MagnetostrictionUses the coupling between magnetic and mechanical properties
Change of length when a magnetic material is magnetized
Used for actuation
• First version of SONAR
• Magnetostrictive
actuators (Terfenol)
Magnetoelastic effectChange in the magnetic state when a magnetic material is deformed
σσ
H = 0
σ > 0σ = 0
produces large permeability changes
~
l
H = 0
l + Δl
H
22
Magnetoelastic sensorsThe magnetoelastic effect can be used to measure deformation, force, torsion, etc.
τ
The detection is non-contact, using pick-up coils to measure permeability changes.
“torductor” ABB
23
Magnetoelastic resonance
h(t) = ho sen ωtAn alternating field excites magneto-elastic waves in the material.
At selected frequencies, mechanical resonances builds up. ε, m, v
ωωr
v(t)
m(t)
!r =n⇡
L
sE
⇢
Young modulus E = E(H): the resonance can be tuned
ωr
HoHoHo
h(t) = ho sen ωt
Ho
24
Magnetoelastic resonanceElectronic article surveillance systems (anti-theft tags)
16 Magnetoelastic Sensors
Acousto-magnetic tag
Plastic sleeve
Magnetostrictive element
Bias magnet
Figure 25. Components of an acousto-magnetic EAS tag. The activeelement is a magnetostrictive amorphous ribbon that is free to oscillateinside a plastic sleeve. The bias magnet is a semi-hard magnetic materialthat can be easily magnetized to activate the tag and demagnetized todeactivate it.
of biasing the sensible element and therefore of selectingits resonance frequency. Figure 26 resumes the operation ofthe system: In the activated state, the bias magnet is mag-netized and the sample resonates at 58 kHz, maintainingan oscillation when the exciting pulse stops. To deactivatethe tag, the authorized operator demagnetizes the bias mag-net (by applying a decreasing-amplitude alternating mag-netic field) so the resonant frequency of the material shiftsto near 60 kHz. The sample only experiments forced oscilla-tions during the existence of the pulse and the receiver doesnot detect any signal between pulses.
Acousto-magnetic EAS systems are working very success-fully around the word and has become a mass application formagnetostrictive amorphous materials. The reliable detec-tion of small tags over large distances in very different con-ditions implies a careful engineering of the sensible material[102, 103].
Figure 26. Principle of operation of an acousto-magnetic EAS system. When the bias magnet is magnetized, it produces a magnetic field Ha on themagnetostrictive element, which resonates at a frequency fa (58 kHz): the tag is activated. In demagnetized state, the field over the element is Hdand its resonance frequency is shifted to fd (about 60 kHz), detuned with the exciting signal: the tag is deactivated.
Other promissing field of sensor development based onthe magnetoelastic resonance is the remote detection ofenvironmental parameters, using the changes of the reso-nant frequency in response to physical paremeters that affectthe magnetostrictive element, such as temperature, stress,pressure, etc. The sensing capabilities of this phenomenaare being actively investigated, and has recently reachedimportant achievements mainly by the work of Grimes andcollaborators. Their investigations and results are compiledin the excellent review “Wireless magnetoelastic resonancesensors: A critical review” [104].
To illustrate the principle of operation of the proposeddevices, let us consider the case of temperature detection.The temperature affects many of the variables that deter-mine the value of the resonance frequency. Apart from thephysical dimensions of the magnetostrictive element, the sat-uration magnetostriction coefficient, the Young’s modulus,the saturation magnetization and the magnetic anisotropyconstants are temperature dependent. All these parametersmake the resonance frequency be dependent on tempera-ture, as is deduced from Eqs. (13) and (14). This depen-dence can be evidenced experimentally: Figure 27 shows theresonance frequency of an amorphous ribbon as a functionof the magnetic field for different temperatures [105]. For agiven value of the applied magnetic field, the resonant fre-quency varies with temperature. According to these results,biasing the resonant element at a given magnetic magneticfield (by means of a suitable polarizing magnet), the sensibil-ity to temperature can be adjusted to different values. Thereis even a compensation point at which the temperature doesnot affect the value of the resonance. The change in theresonant frequency as a function of temperature for differ-ent biasing fields, derived from these results, is presentedin Fig. 28. The use of amorphous ribbons with differentcomposition results in different sensibilities and improved
activated de-activated
Magnetoelastic Sensors 15
Figure 23. Principle of operation of a position sensor based on the magnetostrictive delay line principle. Photographs on the right are differentcommercial versions of the sensor. The housing can be adapted to the specific application, even in a flexible mount. Figures courtesy of MTSSystems Corp. [94].
of exploiting this effect to design useful devices. On theone hand, magnetostrictive materials can be used as tagsor labels for Electronic Article Surveillance (EAS) systems.These tag-and-alarm systems are used, for instance, to avoidunauthorized removal of goods. If the label is activated (inthis context, if it manifests magnetoelastic resonance) whenit crosses an adequate interrogation and detection system, itwill trigger an alarm. Of course, for the system to be useful,a simple method to deactivate the tag, that is, to put it outof resonance, must be provided in case the item be legallytaken out. On the other hand, the magnetoelastic resonancecan be used to monitor any parameter that can affect itsbehavior, usually by producing a shift in the resonancefrequency. Both types of applications, that are reviewednext, share the very convenient feature of allowing remotedetection with suitable means, even through a brick wall orround the corner.
Presently, there are four major technologies used for EASsystems: microwave, magnetic or electromagnetic, radio-frequency and acousto-magnetic [100, 101]. Each one hasdifferent advantages and drawbacks and is more convenientfor a specific situation. The last one is based on the mag-netoelastic resonance and will be described here. In anacousto-magnetic system (Fig. 24), a transmitter emits aradio frequency signal of about 58 kHz in pulses at a rateof 50 to 90 pulses per second. These pulses excite the oscil-lations of the magnetostrictive tag that, if in activated state,resonates exactly at that frequency (58 kHz) and emits a sin-gle frequency signal, like a tuning fork, that is detected by areceiver, triggering an alarm.
In fact, the magnetoelastic oscillations at resonance per-dure when the exciting pulse has finished, so the receiveronly “listen” between pulses. This permits to use signalsof low amplitude (reducing electromagnetic contamination)and to avoid interferences, because the detection is made
when the emitter is off. To deactivate the tag it must beplaced in a situation in which the resonance conditions areno longer met. According to Eq. (14), the resonance fre-quency depends on the Young’s modulus of the material.Fortunately, one of the consequences of the magnetoelas-tic coupling is the !E effect, that is, the dependence ofthe Young’s modulus E on the magnetic state of the mate-rial (Section 2.4). Figure 7 on page 63 shows the depen-dence of the resonance frequency on the external magneticfield. Therefore, if the magnetic state of the magnetostrictiveelement is changed, it no longer resonates at 58 kHz. Theoscillations that are induced by the emitter fade very quicklyafter the exciting pulse has finished and the receiver doesnot detect any signal from the tag. Figure 25 shows the dif-ferent parts of an acousto-magnetic label. The external casecontains a plastic sleeve in which the sensible element, astrip of amorphous metal, is free to oscillate. Also includedis a strip of a semi-hard magnetic material that is responsible
Emitter Receiver
Interrogation zone
Interrogationsignal
Responsesignal
Magnetoelastictag
Figure 24. Electronic Article Surveillance (EAS) system.
25
Magnetoelastic resonanceOil viscosity sensor (for predictive maintenance)
η (cSt)
26
Outline• Introduction
• Magnetism basics
• Magnetization process
• Soft magnetic materials
• Magnetic microsensors • Magnetic field sensors
• Electronic compass
• Magneto-impedance sensors
• Magnetoelastic sensors • Magnetoelastic effect
• Magnetoelastic resonance
• Oil viscosity sensor
• Magnetic nanodiscs for cancer therapy • Magneto-mechanical actuation
• Magnetic vortex state
• Sub-100 nm vortex discs
• Nanodiscs in cancer cells
27
Magnetic nanodiscs for cancer therapyMagnetic nanoparticles in bio-medicine
• Magnetic Resonance Imaging (MRI)
• Drug delivery
• Hyperthermia
• …
Iron oxides, chemically produced.
Patterned magnetic particles• Great shape versatility
• Many different compositions
• Excellent reproducibility
• …
Produced by physical methods (vapor deposition, lithography, …)
28
Magneto-mechanical actuation
Kimetal,NatureMaterials9,165–171(2010)
Magneto-mechanical actuation of Permalloy discs with vortex state
29
Magnetic Vortex statePeculiar magnetic behaviour in the nanoscale: no magnetic domains
Vortex state Magnetization process
• Large permeability and magnetization
• Null remanence
high actuation capability
no particle agglomeration
➞
➞
The completion between exchange and magentostatic energy produce different configurations
30
1 µm ∼ 60 nm
intra-cell actuationexternal actuation
Sub-100 nm vortex discsAllow for intra-cellular magneto-mechanical actuation
• Fabrication of sub-100 nm permalloy discs • Magnetic characterization • In-vitro test of magneto-mechanical actuation
31
Sub-100 nm vortex discsFabrication by Hole-mask colloidal lithography
Use charged spheres to create a distribution of holes on a polymer
1) deposit a PMMA layer (spin coating)
2) charge surface with PDDA+ + +
+ + +- - -
3) deposit charged spheres
non-regular dense arrangement of nanospheres
SiPMMA
32
4) Ti sputtering
PMMASi
6) Oxygen plasma PMMA etching
5) Tape stripping
Ti template of holes
Sub-100 nm vortex discsFabrication by Hole-mask colloidal lithography
33
7) Py sputtering
8) PMMA removal
Py nanodots on Si substrate
Sub-100 nm vortex discsFabrication by Hole-mask colloidal lithography
34
Sub-100 nm vortex discsDetachment from the substrate
Use of Germanium as sacrificial layer
∅ 60 nm
∅ 140 nm
35
-1.0
-0.5
0.0
0.5
1.0
-120 -80 -40 0 40 80 120
M/MsBB
M/M
s
µ0H(mT)
T=50nmR=70nm
Sub-100 nm vortex discsMagnetic behaviour
59
Additional MFM images were recorded under in-plane magnetic fields to
monitor the displacement of the vortex core towards the edge of the disc. The images
nicely correspond to the magnetic states revealed in the hysteresis loop as shown in
Figure 2.13. At zero applied field, the vortex cores are centred, while the core moves
towards the edge of the disc when a small field is applied, being the vortex core
displacement perpendicular to the field direction. As it can be seen in the loop, the
available fields are not enough to completely expel the vortex and go to a saturated
in‐plane configuration.
Figure 2.13. SQUID hysteresis loop of sample L50, with the imaged points marked by red dots and the corresponding MFM images under different in situ fields.
To complement the experimental results, we performed micromagnetic
simulations in discs of the same geometries. Again, we used OOMMF software and
the material parameters described previously except the cell size. Although, the
usual cell size in this kind of simulations is 4 nm, we obtained the same results with
smaller cell sizes. To increase the density of points in Figure 2.14, the data were
obtained using cell sizes of 1 nm for samples S30 and S50 (R = 30 nm), and of 2 nm
for L30 and L50 (R = 50 nm).
Particularly, we were interested in obtaining the configuration of the
magnetisation in the ground state, at zero applied field, to analyse the structure of
36
Sub-100 nm vortex discsMagneto-mechanical actuation
70
Figure 2.19. Light-transmission experiment to study the mechanical responsiveness of the discs to an external magnetic field.
The experiment consists in applying an intermittent magnetic field (tON and
toff) of 2 mT and 1 Hz and a continuous laser beam passing through the aqueous
solution with discs, while the transmitted light intensity is being recorded. The
magnetic field and the laser beam are in the same direction. The resulting plots,
displayed in Figure 2.20, represent the transmitted light intensity and the applied
magnetic field amplitude as a function of time. The dashed line represents the
current in the Helmholtz coils, i.e., the magnetic field, being the tON period
highlighted with pink bars.
The response of the nanodiscs and the microdiscs is similar: the transmitted
light reaches the maximum intensity when the magnetic field is on, and drops when
switched off. This result can be interpreted as the alignment of the plane of the discs
with the magnetic field, which allows the light to pass through the aqueous solution
and reach the detector, as schematically described in Figure 2.21a. However, the
relaxation time (tR) of the microdiscs is clearly larger than the nanodiscs’. We
37
Nanodiscs in cancer cellsInteraction of microdiscs (R = 1 !m, T = 60 nm)nanodiscs (R = 70 nm, T = 50 nm) with human lung carcinoma cells { 86
Figure 3.1. Protocol of the in vitro assays, from the preparation of the cells (steps 1-4) and the treatment (steps 5 and 6) to the cell viability assessment (step 7). CGM: cell growth medium, SN: supernatant.
All of the assays described hereafter, were performed testing the microdiscs
first because, as they have already been studied, there are some reference conditions
in the literature that are helpful to initiate the experiments.
3.2. Intracellular uptake of discs
To study the internalization of the discs by the cells, images were captured by
fluorescence/brightfield microscopy and, more accurately, by transmission electron
microscopy (TEM). Additionally, we recorded live cell videos over 48 h to monitor
Protocol of the in-vitro assays
38
Nanodiscs in cancer cells
86
Figure 3.1. Protocol of the in vitro assays, from the preparation of the cells (steps 1-4) and the treatment (steps 5 and 6) to the cell viability assessment (step 7). CGM: cell growth medium, SN: supernatant.
All of the assays described hereafter, were performed testing the microdiscs
first because, as they have already been studied, there are some reference conditions
in the literature that are helpful to initiate the experiments.
3.2. Intracellular uptake of discs
To study the internalization of the discs by the cells, images were captured by
fluorescence/brightfield microscopy and, more accurately, by transmission electron
microscopy (TEM). Additionally, we recorded live cell videos over 48 h to monitor
Asses the effect of the discs and the alternating magnetic field
39
Nanodiscs in cancer cells89
Figure 3.4. Micrographs of lung carcinoma cells after 24 h of incubation with (a) microdiscs (R = 1 m and T = 60 nm, covered with gold) and (b) nanodiscs (R = 70 nm and T = 50 nm, covered with gold). The discs were added in nominal proportions of 10 and 2000 particles per cell, respectively.
Regarding the internalization mechanism, the process starts at the
interaction between the particles and the cytoplasmic membrane, possibly through
membrane proteins. Proteins contain a wide range of functional groups, including
alcohols, thiols, carboxylic acids, carboxamides and a variety of basic groups able to
react with the gold surface of the discs. Our hypothesis is that the cells internalize
the particles by endocytosis and are subsequently accumulated into lysosomes,
which are specialized organelles that contain hydrolytic enzymes. Lysosomes
function as the digestive system of cells by processing compounds that enter the cell
from the outside, as well as compounds inside the cell.
Transmission electron microscopy (TEM) was performed to provide insight
to the intracellular localization of the discs. After 24 h of incubation with the
particles, the cells were embedded in an epoxy resin and the solidified sample was
cut in 70-90 nm thick slices (the protocol of the sample preparation is described in
Appendix A). As shown in Figure 3.5, microdiscs are localized inside the cell and
oriented perpendicular to the axis, suggesting that they may have been exposed to a
low magnetic field during the fixation process (the same phenomena was observed
in Ref. [12]). Clearly, the area surrounding the microdiscs contrasts with the texture
of the cytoplasm, indicating they could be encapsulated in a lysosome (the holes are
due to the dragging of the microdiscs during the cutting process).
89
Figure 3.4. Micrographs of lung carcinoma cells after 24 h of incubation with (a) microdiscs (R = 1 m and T = 60 nm, covered with gold) and (b) nanodiscs (R = 70 nm and T = 50 nm, covered with gold). The discs were added in nominal proportions of 10 and 2000 particles per cell, respectively.
Regarding the internalization mechanism, the process starts at the
interaction between the particles and the cytoplasmic membrane, possibly through
membrane proteins. Proteins contain a wide range of functional groups, including
alcohols, thiols, carboxylic acids, carboxamides and a variety of basic groups able to
react with the gold surface of the discs. Our hypothesis is that the cells internalize
the particles by endocytosis and are subsequently accumulated into lysosomes,
which are specialized organelles that contain hydrolytic enzymes. Lysosomes
function as the digestive system of cells by processing compounds that enter the cell
from the outside, as well as compounds inside the cell.
Transmission electron microscopy (TEM) was performed to provide insight
to the intracellular localization of the discs. After 24 h of incubation with the
particles, the cells were embedded in an epoxy resin and the solidified sample was
cut in 70-90 nm thick slices (the protocol of the sample preparation is described in
Appendix A). As shown in Figure 3.5, microdiscs are localized inside the cell and
oriented perpendicular to the axis, suggesting that they may have been exposed to a
low magnetic field during the fixation process (the same phenomena was observed
in Ref. [12]). Clearly, the area surrounding the microdiscs contrasts with the texture
of the cytoplasm, indicating they could be encapsulated in a lysosome (the holes are
due to the dragging of the microdiscs during the cutting process).
Even without functionalization, discs are internalized by the cells
∅ 140 nm
2 µm ∅
∅ 2 !m
40
91
For the further experiments, we used a larger concentration of microdiscs
(nominally 25 microdiscs/cell) to increase the population of cells with particles to
that observed using nanodiscs, i.e., 17 %.
3.3. Cytotoxicity
Prior to the study of the destructive capability of the discs under an external
magnetic field, it is essential to evaluate their cytotoxic effect on the lung carcinoma
cells. For that purpose, we followed the protocol described in section 3.1 and tested
the impact of the microdiscs and the nanodiscs on the vital functions of the cells by
adding, respectively, a nominal proportion of 25 and 2000 discs per cell.
Figure 3.7 collects two representative examples of the cytotoxicity tests. As
described in the point 7 of the protocol, the NucBlue dyes the nuclei of all the cells
blue (Figures 3.7b and 3.7e), which allows us to count the total population when
using the DAPI standard filter. The PI instead, only penetrates in dead cells, thanks
to the lower permeability of the dead cells membrane, and dyes the nuclei red, which
can be observed when using the TRITC standard filter (Figures 3.7c and 3.7f).
Representatively, none of the dead cells, marked with white arrows, have discs as
point the red arrows in the brightfield images (Figures 3.7a and 3.7b). We followed
this procedure in the rest of the assays.
Figure 3.7. Micrographs of lung carcinoma cells after 24 h of incubation with microdiscs (R = 1 m and T = 60 nm, covered with gold) and nanodiscs (R = 70 nm and T = 50 nm, covered with gold). (a, d) Microdiscs and the nanodiscs internalized by the cells. (b, e) The nuclei of all the cells. (c, f) Dead cells (marked with white arrows).
After 24 h incubation, nearly 100% of cells with discs survival
Nanodiscs in cancer cellsCytotoxicity
41
94
Figure 3.10. Live cell video captures at different times. At t = 2 h, the cell marked in red divides. At t = 21 h 30 min, it internalizes a group of microdiscs (orange circle). At t = 34 h 30 min, the cell marked in blue divides, whereas the cell with discs is not able to divide and dies after 42 h of incubation.
3.4. Magneto-mechanical stimulus
The last part of the Thesis addresses the study of the efficacy of the magneto-
mechanically actuated nanodiscs to destroy cancer cells. The magnetic field station
was built based on that reported by D-H Kim et al. [9] and is shown in Figure 3.11a.
The AC magnetic field is generated by pair of Helmholtz coils and the well-plate with
the cells is placed at the centre of the station. The magnetic field amplitude was set
to 10 mT with a frequency of 10 Hz; it was reported to be sufficient to dramatically
damage glioma cancer cell using Permalloy discs with R = 0.5 m in [9].
Figure 3.11. Magnetic field station. Helmholtz coils generate an AC field of 10 mT and 10 Hz (a) parallel or (b) perpendicular to the plane of the coverslips as the red arrows indicate. The well-plate is placed in the centre of the air gap as shown in the inset of the figure b.
94
Figure 3.10. Live cell video captures at different times. At t = 2 h, the cell marked in red divides. At t = 21 h 30 min, it internalizes a group of microdiscs (orange circle). At t = 34 h 30 min, the cell marked in blue divides, whereas the cell with discs is not able to divide and dies after 42 h of incubation.
3.4. Magneto-mechanical stimulus
The last part of the Thesis addresses the study of the efficacy of the magneto-
mechanically actuated nanodiscs to destroy cancer cells. The magnetic field station
was built based on that reported by D-H Kim et al. [9] and is shown in Figure 3.11a.
The AC magnetic field is generated by pair of Helmholtz coils and the well-plate with
the cells is placed at the centre of the station. The magnetic field amplitude was set
to 10 mT with a frequency of 10 Hz; it was reported to be sufficient to dramatically
damage glioma cancer cell using Permalloy discs with R = 0.5 m in [9].
Figure 3.11. Magnetic field station. Helmholtz coils generate an AC field of 10 mT and 10 Hz (a) parallel or (b) perpendicular to the plane of the coverslips as the red arrows indicate. The well-plate is placed in the centre of the air gap as shown in the inset of the figure b.
97
For this reason, the magnetic field station was modified in such a way that the field
direction was perpendicular to the plane of the coverslips, as shown in Figure 3.11b.
Furthermore, the perpendicular configuration increases the magnetic flux density
affecting a cell since, as the A549 cells grow in the XY plane, more magnetic field
lines will cross the cell. Both configurations are schematized in Figure 3.14.
Figure 3.14. a) Cells adhered to the coverslip where most of the microdiscs are horizontally oriented. b) In the parallel configuration (Figure 3.11a), when the field is ON, the discs are already aligned to the field. c) In the perpendicular set-up (Figure 3.11b) the angle between the disc plane and the field direction is larger, enhancing the torque of the particle and the magnetic flux density affecting the cell is also higher (magnetic field lines in red).
Then, in the third assay, the magnetic field was applied perpendicular to the
coverslips plane for 30 min. However, the dead cells rate did not increase from 15 %
(the destruction of some cells is shown in Figure 3.15) indicating that the direction
of the magnetic field is neither the key parameter. The other experimental
conditions we could vary were the amplitude and the frequency of the magnetic
field. As it was demonstrated in Chapter 2, a field as small as 2 mT and 1 Hz was
proved sufficient to cause the mechanical torque of the microdisc in water. However,
the movement of the particles in the cytoplasm is probably restricted and a higher
magnetic field may be required. The field applied in the assays is five times larger
(10 mT and 10 Hz) than that used in the light-transmission experiment, which may
be sufficient to cause the rotation of the discs, but once the they are aligned with the
field direction, the particles could get blocked due to the viscosity of the cytoplasm.
Since our magnetic field set-up cannot produce greater magnetic fields, we did not
97
For this reason, the magnetic field station was modified in such a way that the field
direction was perpendicular to the plane of the coverslips, as shown in Figure 3.11b.
Furthermore, the perpendicular configuration increases the magnetic flux density
affecting a cell since, as the A549 cells grow in the XY plane, more magnetic field
lines will cross the cell. Both configurations are schematized in Figure 3.14.
Figure 3.14. a) Cells adhered to the coverslip where most of the microdiscs are horizontally oriented. b) In the parallel configuration (Figure 3.11a), when the field is ON, the discs are already aligned to the field. c) In the perpendicular set-up (Figure 3.11b) the angle between the disc plane and the field direction is larger, enhancing the torque of the particle and the magnetic flux density affecting the cell is also higher (magnetic field lines in red).
Then, in the third assay, the magnetic field was applied perpendicular to the
coverslips plane for 30 min. However, the dead cells rate did not increase from 15 %
(the destruction of some cells is shown in Figure 3.15) indicating that the direction
of the magnetic field is neither the key parameter. The other experimental
conditions we could vary were the amplitude and the frequency of the magnetic
field. As it was demonstrated in Chapter 2, a field as small as 2 mT and 1 Hz was
proved sufficient to cause the mechanical torque of the microdisc in water. However,
the movement of the particles in the cytoplasm is probably restricted and a higher
magnetic field may be required. The field applied in the assays is five times larger
(10 mT and 10 Hz) than that used in the light-transmission experiment, which may
be sufficient to cause the rotation of the discs, but once the they are aligned with the
field direction, the particles could get blocked due to the viscosity of the cytoplasm.
Since our magnetic field set-up cannot produce greater magnetic fields, we did not
H = 10 mTf = 10 Hz
t = 10, 30 min
Nanodiscs in cancer cellsMagneto-mechanical actuation
42
Nanodiscs in cancer cellsMagneto-mechanical treatment
99
50
60
70
80
90
Cel
l via
bilit
y (%
)
MDs MF || 10 min
MDs MF || 30 min
MDsMF 30 min
NanodiscsMF 30 min
Figure 3.16. Percentages of lung carcinoma cell viability (all of them hold microdiscs or nanodiscs) 4h after the application of the magnetic field of 10 mT and 10 Hz.
Figure 3.17. Micrographs of lung carcinoma cells with nanodiscs (R = 70 nm and T = 50 nm, covered with gold), 4 h after the application of the magnetic field of 10 mT and 10 Hz for 30 min, in the perpendicular configuration. In both examples, the cells that have nanodiscs (marked in red in a and d) have been destroyed (red nuclei in c and f).
To explain the effect of the treatment from the biological point of view, a cell
death mechanism induced by the nanodiscs is proposed and summarized in
Figure 3.18, where the lysosomal membrane rupture is suggested to be the principal
cause. P. Saftig et al. have reported that lysosomal membrane permeabilization can
induce the leakage of lysosomal hydrolases into the cytosol, and lead to cell death
eventually [14, 15]. The membrane of the lysosome is a phospholipidic bilayer as it
Cells with nanodiscs inside die after 30 min in perpendicular field
43
99
50
60
70
80
90C
ell v
iabi
lity
(%)
MDs MF || 10 min
MDs MF || 30 min
MDsMF 30 min
NanodiscsMF 30 min
Figure 3.16. Percentages of lung carcinoma cell viability (all of them hold microdiscs or nanodiscs) 4h after the application of the magnetic field of 10 mT and 10 Hz.
Figure 3.17. Micrographs of lung carcinoma cells with nanodiscs (R = 70 nm and T = 50 nm, covered with gold), 4 h after the application of the magnetic field of 10 mT and 10 Hz for 30 min, in the perpendicular configuration. In both examples, the cells that have nanodiscs (marked in red in a and d) have been destroyed (red nuclei in c and f).
To explain the effect of the treatment from the biological point of view, a cell
death mechanism induced by the nanodiscs is proposed and summarized in
Figure 3.18, where the lysosomal membrane rupture is suggested to be the principal
cause. P. Saftig et al. have reported that lysosomal membrane permeabilization can
induce the leakage of lysosomal hydrolases into the cytosol, and lead to cell death
eventually [14, 15]. The membrane of the lysosome is a phospholipidic bilayer as it
Comparison of the effectiveness of the mechanical treatment
Nanodiscs are more effective!
Nanodiscs in cancer cellsMagneto-mechanical treatment
44
Summary
• Soft magnetic materials enable classical technologies but many new applications are continuously being developed.
• Large permeability is a key parameter for magnetic field sensors.
• Magnetic properties are highly coupled with other effects in soft magnetic materials. For instance, magneto-elasticity allows several sensing mechanisms.
• Nanotechnology largely benefits from soft magnetic materials. For example, magneto-mechanical actuation of magnetic nanodiscs with vortex state is studied for novel cancer therapies.
45
Grupo de Magnetismo y Materiales Magnéticos
Acknowledgements