Patterned and switchable surfaces for biomaterial applications

Chapter 5 – Surface plasmon resonance imaging of polymer microarraysBefore spotting polymer samples were prepared to 2, 1, 0.5, 0.25, 0.1, 0.05 and0.01 mg/ml solutions in ultra pure water. A solution of pure water was also spotted asa negative control. For studies with switchable polymers, PNIPAAm was spottedonto gold coated SPR chips at a concentration of 1.5, 0.75, 0.37 and 0.18 mg/ml inwater. Spotting was conducted using a BioOdyssey Calligrapher MiniArrayer (Bio-Rad) using a 375 m diameter solid pin (ArrayIt), a humidity of 65% and at atemperature of 25 °C. The approach speed of the pin and the dwell time of the pin incontact with the surface were set to 20 mm/s and 15 ms respectively. All spotsformed were reprinted thrice directly after initial spot formation to minimisevariation in spot formation.For formation of covalently crosslinked PEI and PLL polymer spots ((PEIc) and(PLLc) respectively), PEI and PLL were spotted onto PEG-modified SPR chips.Before spotting, polymer samples were prepared to 50 l of 0.5 mg/ml solutions inultra pure water containing 1.0 mg/ml (3.3 mM) N-succinimidyl-5-azido-2-nitrobenzoate (NSANB) (Fluka). Polymer-NSANB solutions were incubated at 25°C for 10 min before array formation. For the formation of covalently crosslinkedPAA spots (PAAc), PAA (4.0 mg/ml) was prepared in 50 l of ultra pure watercontaining 75 mM N-hydroxy succinimide (NHS) (Sigma) and 30 mM 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) (Fluka) and incubated for1 hr at 37 °C. 50 l of 2.00 mg/mL (6.6 mM) NSANB in ultra pure water containing7.0 mM ethylenediamine (Merck) at 25 °C was incubated for 10 min before addingto the 50 l of activated PAA, to make a final volume of 100 l, and incubating for afurther 10 min before array formation. After array formation, samples were exposedto UV irradiation (25 W) for at least 10 mins, which was sufficient time to ensure5-164