Biomedical Engineering Reference
In-Depth Information
6.4.2.2
AFM
Sample preparation for force measurements.
Single cellulose beads were glued (epoxy
glue, UHU
) to the end of a tipless cantilever (silicon nitride, Digital Instruments/Veeco,
a nominal spring constant 0.12 N m
−
1
) as described in Paananen
et al
. (2003). For
obtaining more quantitative data, each cantilever was calibrated separately during sample
preparation. The thermal method (Hutter and Bechhoefer 1993) was used for determining
the spring constant. It was chosen for its simplicity, applicability to V-shaped cantilevers
and nondestructive nature. Cellulose beads were attached onto the glass sample support
on a thin layer of glue. Cantilevers and sample supports with cellulose beads were
dried in a desiccator and prepared prior to each force measurement.
+
Also reference
(1 mM NaCl, pH 10) and xylan solutions (100 mg ml
−
1
in 1 mM NaCl, pH 10) were
freshly made.
Force measurements.
The force measurements were done by the colloidal probe tech-
nique (Ducker
et al
. 1991) using a NanoScope IIIa Multimode AFM (Digital Instruments
(Veeco, CA) equipped with a scanner E with vertical engagement, and using an O-ring.
Swelling of the cellulose beads and behavior of the soft cellulose surfaces during force
measurements were studied separately (Paananen
et al
. 2003) in order to perform proper
experiments and interpret the results. Based on these results the cellulose beads were
allowed to swell in water overnight and to stabilize in reference solution (1 mM NaCl,
pH 10) for 2 h prior to the force measurements. Softness of the cellulose surfaces also
required recording force curves with different loading forces. Force curves taken with
low loading force (here
0.6 mN m
−
1
) gave more realistic results for the 'true' interac-
tions than curves of high loading force (here
∼
1.7 mN m
−
1
), but part of the repulsion
measured on approach was due to compression of the cellulose beads and the region of
constant compliance was not reached. Hence, the cantilever sensitivity values needed
for analyzing the force curves with low loading force were taken from the ones with
high loading force, where the constant compliance was reached.
The interaction forces between cellulose surfaces with different loading forces were
measured in a reference solution (1 mM NaCl, pH 10) and in the presence of xylan
(100 mg ml
−
1
in 1 mM NaCl, pH 10). The total exposure time to xylan solution was
5 h before force measurements. All measurements were performed in pH 10 to ensure
that xylan was soluble. The force curves were recorded at slightly different spots still
being on the central area of the beads. The time gap between consecutive force curves
was varied from 0.5 min to 10 min for obtaining information of the relaxation time of
the cellulose surfaces. The measurements were performed several times and the trend
observed in the results was repeatable.
The effect of electrolyte concentration on the interaction between xylan-coated cel-
lulose surfaces (Paananen 2007) was studied by measuring interaction forces between
cellulose beads in 100 mg ml
−
1
xylan solution, pH 10 with varying electrolyte concentra-
tions (1, 10 and 100 mM NaCl). After changing the solution in the measurement cham-
ber, the system was allowed to stabilize for 3 h. The force curves were recorded using
different loading forces and the time gap between consecutive force curves was 5 min.
∼
Analysis of the force curves.
The raw force curve data were converted into ASCII format
using a Scanning Probe Image Processor (SPIP, Image Metrology, Denmark) and further
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