Biomedical Engineering Reference
In-Depth Information
were purchased from Sigma-Aldrich (France). Bis(sulfosuccinimidyl) suberate (BS) was
purchased from Pierce (Rockford, IL, USA).
2.2 Stainless steel surface preparation
The samples (both faces and perimeter) were polished with 1 µm diamond suspension
(Struers, Denmark), rinsed in binary mixture of milliQ water/ethanol (1/1, v/v) in a
sonication bath (70W, 40 kHz, Branson, USA) and dried under nitrogen gas flow.The
samples were then immediately immersed for 48 h in synthetic aqueous medium (NaCl 0.46
mmol.L
-1
, Na
2
SO
4
0.26 mmol.L
-1
, NaNO
3
0.2 mmol.L
-1
, NaHCO
3
3.15 mmol.L
-1
, pH about 8),
abundantly rinsed with milliQ water (Millipore, Molsheim, France) and dried under
nitrogen gas flow. These samples are considered as native SS and designated as “nat”.
2.3 Surface treatment procedure
The silanization of nat samples was performed in the gas phase with APTES. To this end,
nat samples were placed, together with a small vial containing 100 µL APTES, in a 7 dm
3
closed recipient under vacuum for 30 min at room temperature. The samples were cured for
1 h at 100°C under vacuum, rinsed and incubated for 6 h in milliQ water to eliminate the
excess of non attached silanes. These are called “sil”.
Both nat and sil samples were subjected to treatments with the coupling agent (BS) and/or
the enzyme (Gox) as detailed below:
i.
“+BS” and “sil+BS” obtained after BS treatment (10 mM in milliQ water) for 30 min on
nat and sil samples, respectively,
ii.
“+Gox” and “sil+Gox” obtained after Gox treatment (0.1 mg.mL
-1
in phosphate buffer
pH~6.8) for 2 h, on nat and sil samples, respectively,
iii.
“+BS+Gox” and “sil+BS+Gox” obtained after both BS and Gox treatment (according to
the procedure described above) on nat and sil samples, respectively.
After BS or Gox treatment, the samples were rinsed three times with milliQ water and dried
under nitrogen gas flow.
In order to clarify the issue of surface contamination, nat samples were further cleaned by
UV-ozone treatment (UVO Cleaner, Jelight Co, Irvine, Ca, USA) during 20 minutes. They
were then placed in a Petri dish, left in the laboratory environment, and submitted to water
contact angle measurements as a function of time.
2.4 X-ray photoelectron spectroscopy
XPS analyses were performed using a Kratos Axis Ultra spectrometer (Kratos Analytical,
UK), equipped with a monochromatized aluminum X-ray source (powered at 10 mA and 15
kV) and an eight channeltrons detector. No charge stabilization device was used on these
conductive samples. Analyses were performed in the hybrid lens mode with the slot
aperture; the resulting analyzed area was 700 µm
¯
300 µm. A pass energy of 40 eV was
used for narrow scans. In these conditions, the full width at half maximum (FWHM) of the
Ag 3d
5/2
peak of clean silver reference sample was about 0.9 eV. The samples were fixed on
the support using a double-sided adhesive conducting tape. The pressure in the analysis
chamber was around 10
-6
Pa. The photoelectron collection angle
θ
between the normal to the
sample surface and the electrostatic lens axis was 0° or 60°. The following sequence of
spectra was recorded: survey spectrum, C 1s, O 1s, N 1s, P 2p, Cr 2p, Fe 2p, Ni 2p, Mo 3d,
Na 1s, S 2p, Si 2p and C 1s again to check for charge stability as a function of time, and
absence of sample degradation. The binding energy scale was set by fixing the C 1s
