A. STAMBOLI] et al.: DETERMINATION OF MECHANICAL AND FUNCTIONAL PROPERTIES BY CONTINUOUS ...
521–527
DETERMINATION OF MECHANICAL AND FUNCTIONAL
PROPERTIES BY CONTINUOUS VERTICAL CAST NiTi ROD
DOLO^ITEV MEHANSKIH IN FUNKCIONALNIH LASTNOSTI
VERTIKALNO KONTINUIRNO LITE NiTi PALICE
Ale{ Stamboli}
1,2
, Monika Jenko
1,2
, Aleksandra Kocijan
1
, Borut @u`ek
1
, Damjana Drobne
3
,
Rebeka Rudolf
4,5
1
Institute of Metals and Technology, Lepi pot 11, 1000 Ljubljana, Slovenia
2
Jo`ef Stefan International Postgraduate School, Jamova cesta 39, 1000 Ljubljana, Slovenia
3
Biotechnical Faculty, Department of Biology, Ve~na pot 111, 1000 Ljubljana, Slovenia
4
University of Maribor, Faculty of Mechanical Engineering, Smetanova ulica 17, 2000 Maribor, Slovenia
5
Zlatarna Celje d.o.o., Kersnikova ulica 19, 3000 Celje, Slovenia
ales.stambolic@imt.si
Prejem rokopisa – received: 2018-02-26; sprejem za objavo – accepted for publication: 2018-08-23
doi:10.17222/mit.2018.030
This article presents the information of the microstructure and methods for determination of mechanical and functional proper-
ties of the Continuous Vertical Cast (CVC) NiTi rod. We prepared special samples (discs, cylinders). The results of the micro-
hardness measurements with HV0.1 show the average Vickers hardness value 583 HV0.1 in the cross-section of the CVC NiTi
rod measured from the edge toward the middle of CVC NiTi rod. The results show that Vickers hardness slightly increases
towards the center of the rod as a consequence of the formatted microstructure. The results of the tensile test show a typical
curve for brittle materials. The obtained results indicate that the CVC NiTi rod does not pass into a non-linear plastic region.
Therefore, CVC NiTi rod is unsuitable for strain hardening. The results of the compressive test also follow the conclusions that
CVC NiTi rod is brittle material. Fracture of the CVC NiTi rod sample occurs before the material enters the plastic deformation
region. The cytocompatibility of the CVC NiTi rod was tested with the submersion of the samples to the HUVEC cell solution,
where HUVEC cells adhered to the samples surface. A commercial NiTi alloy was used to monitor the cytocompatibility. The
results show that the number of adhered HUVEC cells on the samples surface of CVC NiTi rod is smaller than on commercial
NiTi. The number of dead HUVEC cells on the samples surface of CVC NiTi rod is slightly larger than on commercial NiTi.
The proportion of viable HUVEC cells on the samples surface of CVC NiTi rod is 96,6 %, while on commercial NiTi 98,4 %.
Keywords: CVC NiTi rod, hardness, mechanical properties, cytocompatibility
V ~lanku je podana informacija o mikrostrukturi in metodah za dolo~itev mehanskih in funkcionalnih lastnosti vertikalno-konti-
nuirno lite (CVC) NiTi palice. Pripravljeni so bili posebni vzorci (diski, valji). Rezultati meritev mikrotrdote HV0.1 ka`ejo
povpre~no vrednost trdote v pre~nem prerezu CVC NiTi palice, merjeno od roba proti sredini, 583 HV0.1. Vrednost trdote rahlo
nara{~a proti sredini palice in je posledica nastale mikrostrukture. Rezultati nateznega preizkusa ka`ejo zna~ilno krivuljo za
krhke materiale in nakazujejo, da CVC NiTi palica ne preide v nelinearno plasti~no obmo~je. Zaradi tega je CVC NiTi palica
neprimerna za deformacijsko utrjevanje. Tudi rezultati tla~nega preizkusa sledijo ugotovitvam, da CVC NiTi palica spada med
krhke materiale. Testirali smo citokompatibilnost CVC NiTi palice z izpostavljenostjo vzorcev raztopini HUVEC celic, kjer so
se le-te adhezirale na povr{ini vzorcev. HUVEC celice so tipi~ne `ilne celice, nahajajo se predvsem v venah. Za kontrolni
material pri citokompatibilnosti je bila uporabljena komercialna NiTi zlitina. Rezultati ka`ejo, da je {tevilo adheziranih celic na
povr{ini vzorca CVC NiTi palice nekoliko ni`ja kot na komercialni NiTi zlitini. [tevilo mrtvih HUVEC celic na povr{ini vzorca
CVC NiTi palice je nekoliko vi{ja kot na komercialni NiTi zlitini. Dele` `ivih celic na povr{ini vzorca CVC NiTi palice je
96,6 %, medtem ko je na komercialni NiTi 98,4 %.
Klju~ne besede: CVC NiTi palica, trdota, mehanske lastnosti, citokompatibilnost
1 INTRODUCTION
Nitinol is a group of nearly equi-atomic alloys com-
posed of nickel and titanium. It exhibits a unique
combination of good functional properties and a high
mechanical strength, such as superelasticity and a
shape-memory effect, good corrosion resistance in a
chloride environment, an unusual combination of
strength and ductility, high tendency for self-passivation,
paramagnetic properties, high fatigue strength, low
Young’s modulus and excellent bio-mechanical compa-
tibility.
1–7
This alloy was developed in the 1970s and its pro-
perties have enabled its use, especially for biomedical
purposes, first in orthodontic treatments, and later on in
cardiovascular surgery for stents, guide wires, filters,
etc., in orthopedic surgery for various staples and rods,
and in maxillofacial and reconstructive surgery. In addi-
tion to bioengineering, nitinol has been used in aero-
space, automotive, marine, chemical industries, civil and
structural engineering.
1–7
Biocompatible materials can be defined as synthetic
or natural biomaterials that are continuously or occasion-
ally in contact with body fluids and can partially or
totally replace any failing tissue, organ or function of the
body in order to improve the quality of life of the indi-
vidual. To successfully employ implants in the human
body, a suitable level of tolerance of the material used
Materiali in tehnologije / Materials and technology 52 (2018) 5, 521–527 521
UDK 67.017:669-147:669.24'295 ISSN 1580-2949
Original scientific article/Izvirni znanstveni ~lanek MTAEC9, 52(5)521(2018)

within the living organism is required.
8–10
The main
problem of nitinol’s biocompatibility is chemical com-
position and consequently high Ni content. Ni releasing
can induce toxic, allergic and hypersensitive reactions or
tissue necrosis after long term implantation. Leaching of
Ni can arise when a strongly acidic fluid attacks the
surface of the alloy. This corrosion is accompanied by
nickel release from an implant into the surrounding body
fluid and tissue, which can enhance an allergic reaction
in a sensitive organism.
Second, corrosion can cause pitting and a reduction
of mechanical performance of an implant. In an extreme
case, an implant broken due to corrosion might produce
sharp fragments that are dangerous for the surrounding
tissue. To prevent corrosion and consequently Ni release,
a coating of appropriate thickness must be formed on the
NiTi surface. Titanium oxide coatings are useful enough
to suppress nickel ions outleaching.
11–15
The production of CVC NiTi rod was discussed in
our previous paper.
16
In this article we present the
methods and approaches for determination of hardness
and mechanical properties of as-cast CVC NiTi rod (as
tensile, compressive and bending tests). Further, the
comparison of CVC NiTi rod cytocompatibility with the
commercially available nitinol in the form of rolled sheet
was tested.
2 EXPERIMENTAL PART
2.1 Sample preparation
For microstructure investigation and cytocompati-
bility testing of the CVC NiTi rod was cut transversally
in the disc shape with the diameter of the rod (approx-
imately 11 mm) and a width of 1 cm (Figure 1). For this
purpose, an Accutom 50 (IMT, Ljubljana, Slovenia)
electronic saw was used for precision cutting. Com-
mercial NiTi, which was in the rolled state in the form of
sheets with width of 1.5 mm, was cut to circles with
diameter of 10 mm and width of 1.5 mm using water jet
cutter. For easier polishing, the samples were then hot-
pressed into Bakelite. Mechanical polishing was per-
formed on Struers Abramin apparatus (IMT, Ljubljana,
Slovenia). The grinding was performed with 320-grit SiC
abrasive paper, mechanical polishing with MD-Largo
discs with 9 μm diamond suspension and with peroxide
grains in a chemically aggressive suspension – OP-S
(colloidal silica). At the end samples were cleaned with
detergent, well washed with water and put in an
ultrasound bath in alcohol. For the microstructure
characterization, samples were then etched with Kroll’s
reagent (3 mL HF, 6 mL HNO3 and 100 mL of distilled
water).
The chemical composition of CVC NiTi rod and
commercial NiTi was measured using Inductively
Coupled Plasma Optical Emission Spectroscopy –
ICP-OES (Agilent 720, IMT, Ljubljana, Slovenia).
Microstructure investigation of NiTi discs was per-
formed using Scanning Electron Microscopy – Thermal
Field Emision SEM JEOL JSM-6500F, equipped with
Energy-dispersive X-ray spectroscopy (EDS), Wave-
length-dispersive X-ray spectroscopy (WDS) and
Electron backscatter diffraction (EBSD) analytical tech-
niques (IMT, Ljubljana, Slovenia).
2.2 Hardness and mechanical properties
The hardness of the CVC NiTi rod was measured by
Vickers. For this purpose, the CVC NiTi rod was cut
transversally. Vickers hardness was measured in multiple
points from the end of the rod to the center (radius) of
the rod. Vickers hardness was performed on Instron
Tukon 2100B (IMT, Ljubljana, Slovenia). The selected
load was HV0.1, which means that a force of 0.98 N was
used.
The tensile test samples were turned according to the
B8 × 40 standard (Figure 2a). The diameter of the spe-
cimens was 8 mm, and their length was 40 mm, and an
M10 thread was used. The compressive test samples
were turned into cylinders with a diameter of 10 mm and
a height of 12 mm (Figure 2b). Tensile and compressive
tests were accomplished at room temperature (20±0.5 °C).
Tests were performed on INSTRON 1255, 500 kN
device (IMT, Ljubljana, Slovenia). Preload force was
A. STAMBOLI] et al.: DETERMINATION OF MECHANICAL AND FUNCTIONAL PROPERTIES BY CONTINUOUS ...
522 Materiali in tehnologije / Materials and technology 52 (2018) 5, 521–527
Figure 2: Schematic presentation of characteristic samples for: a) ten-
sile test, b) compressive test and c) 4-point bending test Figure 1: Disc sample for cytocompatibility measurement

1kN and the velocity of deformation increase was
0.00025 s
–1
in tension and in compression the preload
force was 1 kN and the compression velocity was
2 mm/min.
The 4-point bending test specimens were turned into
cylinders with diameter of 4 mm and length of 12 mm
(Figure 2c). The 4-point bending test was accomplished
at room temperature (19 °C). Tests were performed on
AMSLER device (IMT, Ljubljana, Slovenia). The
preload force was 0.0125 kN and bending velocity was
0.5 mm/min.
2.3 Cytocompatibility
Human Umbilical Vein Endothelial Cells (HUVEC
cells) were cultured in Dulbecco’s modified Eagle’s
medium, in which 4-mM L-glutamine and 10 % (v/v)
foetal bovine serum (FBS) was added. Cells were grown
at 37 °C in a humidified atmosphere with 5 % CO
2
and
were routinely passaged twice a week.
Sterile samples were aseptically put into the 12-well
plates and then the cells were seeded into each well
(2×10
4
cells/cm
2
). After a 24-hour incubation, allowing
the cells to adhere, the cells on the samples were rinsed
with Dulbecco’s Phosphate-Buffered Saline (to remove
any floating, unattached cells) and stained with 2 μg/mL
Hoechst 33342 and 2 μg/mL Propidium iodide for 20
min. Hoechst 33342 stains nuclei of all cells blue, while
Propidium iodide stains nuclei of dead cells red. Stained
cells that were adhered to the samples were observed
with a fluorescent microscope Axio Imager.Z1 (Biotech-
nical Faculty – Department of Biology, Ljubljana, Slo-
venia) immediately after the staining. For each sample,
several representative pictures at 25× and 100× magni-
fications were taken. The pictures were processed by the
use of ImageJ 1.46 r software, where the number of all
attached cells (based on the number of blue nuclei) and
the number of dead cells (based on the number of red
nuclei) were evaluated.
After microscopic observation, samples with attached
cells were inserted into the Karnovsky fixative, consist-
ing of 2.5 % glutaraldehyde and 0.4 % paraformaldehyde
in 1-M Na-phosphate buffer (NaH
2
PO
4
·2H
2
Oa n d
Na
2
HPO
4
·H
2
O). After 3 h incubation at room tempera-
ture, fixative was removed and samples were washed for
3 × 10 min with 1-M Na-phosphate buffer.
The post-fixation of samples was accomplished by
1 % osmium tetroxide (OsO4) for 60 min, followed by
washing in distilled H
2
O (3 × 10 min).
After the cells were fixed, images of cells on the
surface of the samples were recorded with light micro-
scopy Microphot FXA (IMT, Ljubljana, Slovenia). The
proportion of the occupied surface with cells was
measured using optical microscopic images and the
computer software analySIS.
3 RESULTS AND DISCUSSION
3.1 Microstructure of as-cast CVC NiTi rod
Using CVC, a 3 m long rod with diameter of approxi-
mately 11 mm was obtained (Figure 3). The rod was
silver coloured with visible periodic marks on its surface.
ICP-OES analysis for the first attempt of CVC NiTi rod
showed a constant material composition of x
Ni
= 59.8 %,
x
Ti
= 38.9 %, x
C
= 0.3 % and approximately x
Fe
=1%.
The microstructure of our CVC NiTi rod (Figure 4)
is dendritic, with a radial orientation of the grains. The
microstructure changes along the rod radius, and from a
distance from the shell of a rod, it passes from fine-
grained to rough with heavily expanded dendritic
branches and a greater proportion of the interdendritic
eutectic. In the microstructure there are no visible
defects such as cracks and other type of porosity.
Secondary electrons image (Figure 4) shows typical
dendritic structure that is solidified primarily (NiTi
phase). At the eutectic temperature (1118 °C) typical
lamellar eutectic structure (NiTi + TiNi
3
) is solidified
from the residue of the melt. All the phases were con-
firmed using EBSD analysis.
3.2 Microstructure of commercial nitinol
The secondary electrons image of the nitinols cross-
section (Figure 5) reveals a relatively microstructure that
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Materiali in tehnologije / Materials and technology 52 (2018) 5, 521–527 523
Figure 4: Secondary electron image at 2500x magnification of CVC
NiTi rod Figure 3: CVC NiTi rod

is characteristic for materials that were produced by
rolling processes, since it is possible to notice a cha-
racteristic texture. Grains are almost the same size (about
20 μm), grain boundaries are clearly noticeable. Com-
mercial nitinol consist of 2 phases. The EDS analysis
showed that the prevailing phase (dark phase) is NiTi,
containing x
Ni
=50%andx
Ti
= 50 %. The second phase
is carbon and titanium rich phase (bright phase – x
C
=
33.3 %, x
Ti
= 40.1 %, x
Ni
= 26.6 %).
The chemical composition of commercial NiTi,
measured with ICP-OES analysis, is x
Ni
= 50.1 %, x
Ti
=
49.89 % and x
C
< 300 μg/g.
3.3 Hardness
The most basic information about material is hard-
ness. The cooling of CVC NiTi rod during drawing was
held perpendicular to the drawing direction. The lowest
temperature was on the mould walls or in the edge of
CVC NiTi rod and was rising from circumference of
CVC NiTi rod to the middle of CVC NiTi rod. In this
manner we were interested in how the difference in the
cooling temperature between the edge and the middle of
CVC NiTi rod affects the hardness of the CVC NiTi rod.
The hardness was measured in the CVC NiTi rod
cross-section in its radius (from the edge of CVC NiTi
rod to the middle of CVC NiTi rod).
As can be seen from diagram in Figure 6, the hard-
ness of CVC NiTi rod is very constant in the direction of
its radius. On the edge (0) of CVC NiTi rod the hardness
is app. 550 HV and the hardness value is then slightly
increasing and alternating to the middle of the rod (D/2),
where the hardness value is app. 600 HV. It can be
concluded that the difference in temperature during
cooling between the edge and the middle of CVC NiTi
rod does not affect the hardness significantly. As we used
a smaller tip to measure hardness, thus the differences in
measured values of the hardness can be greater as the tip
captures a smaller area.
17
The length of indentation
diagonal of used pyramid tip is about 13 nm long which
means that when the hardness value is smaller, the tip
makes indentation in an area where the dendrite is the
prevailing phase. When the hardness value is larger, the
tip makes indentation in a lamellar area and larger value
is a consequence of harder Ni rich phase (Ni
3
Ti).
Material with hardness 550–600 HV can be assumed
to be very hard. Motemani et al.
18
measured the hardness
of 50.7NiTi to be over 500 HV when it was cooled
rapidly with water, while the same material had hardness
below 400 HV if it was cooled slowly in furnace. Also
higher Ni content means higher hardness.
19
From this it
can be concluded that over 550 HV hardness of CVC
NiTi rod is comprehensible due to its high Ni content (x
Ni
= 60 %) and fast water cooling during drawing.
4 MECHANICAL PROPERTIES
4.1 Tensile test
The picture of test specimens before and after the
tensile test is seen in Figure 7. The fracture of test spe-
cimens occurred in the thread part where the test speci-
mens are the weakest. Also the fracture was typically
brittle (no neck has occurred).
A. STAMBOLI] et al.: DETERMINATION OF MECHANICAL AND FUNCTIONAL PROPERTIES BY CONTINUOUS ...
524 Materiali in tehnologije / Materials and technology 52 (2018) 5, 521–527
Figure 6: Diagram of Vickers hardness for CVC NiTi rod with the
distance r Figure 5: SE image at 2500× magnification of commercial nitinol
Figure 7: A test specimen of CVC NiTi rod before and after the
tensile test

The tensile test diagram (Figure 8) shows a typical
curve for brittle materials. Test 1 has a fracture tension,
 T,u
, of 388 MPa, a fracture strain,  T,u
, of 0.9 %, and an
elastic modulus in tension, E
m,T
, of 40 GPa, while test 2
has a fracture tension of 347 MPa, a fracture strain of
0.8 % and an elastic modulus in tension of 44 GPa. Both
curves have a fairly linear shape up to the fracture. This
means that the entire curves are located in an elastic
region where the deformations are completely recover-
able upon removal of the load. Such a diagram does not
have a non-linear plastic region where deformations
become permanent. If the material is sufficiently ductile,
tensile stress can also greatly increase with high plastic
deformation. CVC NiTi rod does not reach the plastic
deformation and is thus unsuitable for strain hardening.
The obtained results correspond to the literature,
18–20
where the authors determined that the brittleness of the
as-cast NiTi alloy is mainly influenced by the high cool-
ing rate of the alloy and its higher nickel content. If the
as-cast NiTi alloy is furnace cooled or later hot worked,
it also shows ductile properties.
4.2 Compressive test
In contrast to the tensile test, where the specimen
withstands tensile loads and is thus elongated, by the
compressive test the specimen resists to compressive
strength and is thus pushed together. Figure 9 shows the
specimen before and after the compressive test. Before
the test, we had a cylinder-shaped specimen that was
crushed to a large number of small pieces after the
achieved ultimate compressive strength.
The diagram in Figure 10 shows the curves of brittle
material in the compressive test. Test 1 has the ultimate
compressive strength,  C,u
, of 2350 MPa, the ultimate
compressive strain,  C,u
, is 10.5 %, the elastic modulus in
compression, E
m,C
is 29 GPa, and test 2 has the ultimate
compressive strength of 2380 MPa, the ultimate
compressive strain of 11.5 %, and elastic modulus in
compression of 27 GPa. Both curves are very similar,
and again they are located in the elastic area of the
compressive test. Unlike the tensile test, this time there is
a slight plastic plateau above 2000 MPa. The compres-
sive test also shows that the CVC NiTi rod is brittle and
is practically incapable of plastic deformations, and is
therefore unsuitable for strain hardening.
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Materiali in tehnologije / Materials and technology 52 (2018) 5, 521–527 525
Figure 9: A test specimen of CVC NiTi rod before and after com-
pressive test
Figure 8: Stress-strain curve for tensile test of both test specimens of
CVC NiTi rod
Figure 11: Stress-strain curve for 4-point bending test of three test
specimens of CVC NiTi rod
Figure 10: Stress-strain curve for compressive test of both test speci-
mens of CVC NiTi rod

4.3 4-point bending test
The 4-point bending diagram in Figure 11 shows
predominantly linear curves. As expected, the curves are
located in the elastic area. This time the fracture again
occurs before the curves reach the area of plastic
deformation. Test 1 has the ultimate flexure stress,  F,u
,
of 427 MPa, ultimate flexure strain,  F,u
,o f0 . 5%a n d
elastic modulus in flexure, E
m,F
, of 113 GPa. Test 2 has
ultimate flexure stress of 610 MPa, ultimate flexure
strain of 0.6 % and elastic modulus in flexure of 126
GPa. Test 3 has ultimate flexure stress of 520 MPa,
ultimate flexure strain of 0.6 % and elastic modulus in
flexure of 110 GPa.
4.4 Cytocompatibility
For the purpose of cytocompatibility measurement,
the samples were submersed in a HUVEC cells solution,
where the HUVEC cells adhered to the samples surface.
HUVEC cells are typical cells in blood vessels,
especially veins.
Figure 12a shows the diagram of all the adhered
HUVEC cells on the surface of CVC NiTi rod and
commercial NiTi. The number of adhered HUVEC cells
on the surface of the CVC NiTi rod is 3124/mm
2
, and on
commercial NiTi 5524/mm
2
. It can be seen that the
number of HUVEC cells adhered to the surface of the
commercial NiTi is higher than on the surface of the
CVC NiTi rod. In Figure 12b, the number of dead
HUVEC cells is shown. The number of dead HUVEC
cells on the surface of the CVC NiTi rod is 106/mm
2
,
and on commercial NiTi it is 91/mm
2
.
Optical microscope images (Figure 13a,b)o f
HUVEC cells on CVC NiTi rod and commercial NiTi
show that the cells are arranged individually on both
substrates, and they are nowhere grouped together. On
commercial NiTi, the cells are much more elongated,
while on CVC NiTi rod they are much more scrunched
together. In combination with Figure 13c showing a
percentage of covered surface area with HUVEC cells, it
can be seen that more occupied area with HUVEC cells
is on commercial NiTi (23,1 %), than on the CVC NiTi
rod (13,2 %).
Commercial NiTi is more cytocompatible than CVC
NiTi rod. It has more HUVEC cells adhered to its sur-
face, less dead HUVEC cells and higher proportion of
the surface area covered with HUVEC cells. The litera-
ture review
21–25
explains that the growth of cells on the
surface of nitinol is influenced by nickel and oxidation of
the nitinol surface. The higher the concentration of
nickel, the lower are biocompatible properties due to
possibility of releasing of Ni ions that may cause toxic,
allergic, and potentially carcinogenic effects,
21
while
biocompatibility can be improved with oxidation, as the
TiO
2
layer is formed and thus the nickel is removed from
the surface. Commercial NiTi (50Ni50Ti) has better
biocompatibility due to the fact that it contains less
nickel than the CVC NiTi rod (60Ni40Ti).
5 CONCLUSIONS
The hardness measurement of the as-cast CVC NiTi
rod has revealed that it is very hard (> 550 HV), which is
influenced by high cooling rates during CVC process
and higher nickel content.
The results of the tensile, compressive and bending
tests in the form of diagrams evidently showed that the
CVC NiTi rod breaks before it passes from the elastic to
the plastic region. The CVC NiTi rod is therefore brittle
material and cannot be strain hardened.
The biocompatibility test with HUVEC cells showed
that more biocompatible is the commercial NiTi because
it has more HUVEC cells adhered to its surface, less
dead HUVEC cells and higher proportion of the surface
area covered with HUVEC cells than CVC NiTi rod.
A. STAMBOLI] et al.: DETERMINATION OF MECHANICAL AND FUNCTIONAL PROPERTIES BY CONTINUOUS ...
526 Materiali in tehnologije / Materials and technology 52 (2018) 5, 521–527
Figure 13: Optical microscope images of HUVEC cells on a) CVC NiTi rod, b) commercial NiTi and c) covered surface area with HUVEC cells
Figure 12: a) Number of adhered HUVEC cells and b) number of
dead HUVEC cells

Acknowledgements
This work was carried out within the framework of
the P2-0132 and L2-5486 of the Slovenian Research
Agency and the PhD grant no. 10/2013.
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