144
7 Reactive μCP on Ultrathin Block Copolymer Films …
7.5 Experimental Section
Materials: PS 690 -b-PtBA 1210 diblock copolymer was purchased from Polymer
Source Company (Dorval, Canada) and used as received. The molar mass and
polydispersity were 202.4 kg/mol and 1.03, respectively. Amino end-labeled PEG
(denotes as PEG 500 NH 2 ), purchased from Nektar UK Company (M n = 500 g/mol,
PDI = 1.1), fluoresceinamine (Molecular Probes, Inc. The Netherlands), and the
DNA samples (probe DNA: P: 25mer 5
-GGA ATG TGC CAT ACC GAA TCC
GTG T-3
; Cy5-labeled target DNA: 5
-CAC GGA TTC GGC ATG-3
; Cy5-labeled
mismatch DNA: 5
-TGT GCC TAA GCC ATA-3
(MWG BIOTEC AG, Ebersberg,
Germany) were used as received. DNA samples were stored at −4 °C until use.
Bovine serum albumin (BSA) with Alexa Fluor® 594 conjugate was bought from
Molecular Probes and used as received.
Preparation of Thin Films. Thin polymer films were prepared by spin-coating
polymer solutions in toluene (typical concentration between 10 and 20 mg/ml) onto
silicon wafers (111) or glass (cover slide from Menzel-Glaser), which were previously cleaned by an oxygen plasma using an Elektrotech PF 340 apparatus (pressure
of O 2 : 0.5 bar; current: 30 mA). All spin-cast samples were dried at room temperature
for 24 h in vacuum before analysis. Mean film thicknesses were determined using a
custom-built spectroscopic ellipsometry using a He-Ne laser (λ = 632.8 nm). The
refractive index of the PEG films was approximated as 1.4638, while for polymer as
1.513 [25].
Functionalization of Hydrolyzed Polymer Films. The COOH groups obtained on
the polymer films after hydrolysis were activated by immersion into an aqueous
solution of 1-ethyl-3-(dimethylamino)-propylcarbodiimide (EDC, 1 M) and Nhydroxysuccinimide (NHS, 0.2 M) for 30 min. The samples were then rinsed with
Milli-Q water, dried under a stream of nitrogen, and used immediately thereafter.
Local Hydrolysis and Fluoresceinamine Coupling. The PDMS stamps prepared
according to the literature [26] were mildly oxidized in an UV ozone plasma reactor
for 50 min. A drop of trifluoroacetic acid (50 μl) was applied on the stamp (1 × 1 cm
2 )
and the acid was dried in air from 30 s to 4 min. Then the stamp was brought to contact
with the surface of polymer films and kept for 20 min. After peeling off the stamp, the
films were rinsed three times using Milli-Q water and were subsequently activated
with NHS/EDC. The films were immersed in fluoresceinamine solution (100 μM,
PB, pH = 7.4). Then the samples were taken out, rinsed by PB and Milli-Q water,
and dried in a stream of nitrogen.
Homogeneous Hydrolysis and (Bio)molecule Coupling. The polymer films were
hydrolyzed in trifluoroacetic acid liquid phase for 20 min, rinsed three times using
Milli-Q water, and functionalized with NHS/EDC. For PEG 500 NH 2 coupling from
solution, the polymer films were subsequently immersed into the corresponding
PEG 500 NH 2 solution (200 μM PEG 500 NH 2 solution, PB, pH = 7.4). After a reaction
time between 5 min and 10 h, the samples were taken out of the solution and were
7 Reactive μCP on Ultrathin Block Copolymer Films …
7.5 Experimental Section
Materials: PS 690 -b-PtBA 1210 diblock copolymer was purchased from Polymer
Source Company (Dorval, Canada) and used as received. The molar mass and
polydispersity were 202.4 kg/mol and 1.03, respectively. Amino end-labeled PEG
(denotes as PEG 500 NH 2 ), purchased from Nektar UK Company (M n = 500 g/mol,
PDI = 1.1), fluoresceinamine (Molecular Probes, Inc. The Netherlands), and the
DNA samples (probe DNA: P: 25mer 5
-GGA ATG TGC CAT ACC GAA TCC
GTG T-3
; Cy5-labeled target DNA: 5
-CAC GGA TTC GGC ATG-3
; Cy5-labeled
mismatch DNA: 5
-TGT GCC TAA GCC ATA-3
(MWG BIOTEC AG, Ebersberg,
Germany) were used as received. DNA samples were stored at −4 °C until use.
Bovine serum albumin (BSA) with Alexa Fluor® 594 conjugate was bought from
Molecular Probes and used as received.
Preparation of Thin Films. Thin polymer films were prepared by spin-coating
polymer solutions in toluene (typical concentration between 10 and 20 mg/ml) onto
silicon wafers (111) or glass (cover slide from Menzel-Glaser), which were previously cleaned by an oxygen plasma using an Elektrotech PF 340 apparatus (pressure
of O 2 : 0.5 bar; current: 30 mA). All spin-cast samples were dried at room temperature
for 24 h in vacuum before analysis. Mean film thicknesses were determined using a
custom-built spectroscopic ellipsometry using a He-Ne laser (λ = 632.8 nm). The
refractive index of the PEG films was approximated as 1.4638, while for polymer as
1.513 [25].
Functionalization of Hydrolyzed Polymer Films. The COOH groups obtained on
the polymer films after hydrolysis were activated by immersion into an aqueous
solution of 1-ethyl-3-(dimethylamino)-propylcarbodiimide (EDC, 1 M) and Nhydroxysuccinimide (NHS, 0.2 M) for 30 min. The samples were then rinsed with
Milli-Q water, dried under a stream of nitrogen, and used immediately thereafter.
Local Hydrolysis and Fluoresceinamine Coupling. The PDMS stamps prepared
according to the literature [26] were mildly oxidized in an UV ozone plasma reactor
for 50 min. A drop of trifluoroacetic acid (50 μl) was applied on the stamp (1 × 1 cm
2 )
and the acid was dried in air from 30 s to 4 min. Then the stamp was brought to contact
with the surface of polymer films and kept for 20 min. After peeling off the stamp, the
films were rinsed three times using Milli-Q water and were subsequently activated
with NHS/EDC. The films were immersed in fluoresceinamine solution (100 μM,
PB, pH = 7.4). Then the samples were taken out, rinsed by PB and Milli-Q water,
and dried in a stream of nitrogen.
Homogeneous Hydrolysis and (Bio)molecule Coupling. The polymer films were
hydrolyzed in trifluoroacetic acid liquid phase for 20 min, rinsed three times using
Milli-Q water, and functionalized with NHS/EDC. For PEG 500 NH 2 coupling from
solution, the polymer films were subsequently immersed into the corresponding
PEG 500 NH 2 solution (200 μM PEG 500 NH 2 solution, PB, pH = 7.4). After a reaction
time between 5 min and 10 h, the samples were taken out of the solution and were