Patterned and switchable surfaces for biomaterial applications

Chapter 4 – Formation of a chemically patterned substrate for cell microarray applicationssufficient time to ensure complete reaction of the crosslinker. This resulted in theformation of crosslinked PEI and PLL spots ((PEIc) and (PLLc) respectively). In thiscase, PAA and PVP were used as negative controls. The UV lamp (multiband source,254-365 nm) was held 1 cm from the sample. Resultant polymer arrays were washedovernight in 0.05% Tween20 (Aldrich) solution at 37 °C with stirring before rinsingwith MilliQ water and drying under nitrogen. For preparation of a polymer arraywith subsequent DNA array formation, the initial PLLc polymer array was depositedusing a 1000 µm diameter solid pin (ArrayIt). This pin was also used for subsequentspotting of DNA and transfection reagent. After formation of the polymer array thechips were removed from the MiniArrayer, exposed to UV and washed in MilliQwater before being dried under a nitrogen stream and placed back in theMiniArrayer. Positioning blocks within the MiniArrayer were used to ensure exactre-positioning of removed slides. Plasmids pEGFP-N1, encoding for the greenfluorescence protein (GFP), and pDsRed2, encoding for the red fluorescence protein(RFP), were then arrayed onto the polymer spots in a checkerboard pattern at aconcentration of 100 µg/ml in nuclease free water at a humidity of 65% and atemperature of 15 °C. DNA spots were reprinted thrice to increase surface coverage.Effectene transfection agent (QIAGEN) was spotted directly on top of the DNAarray at the same atmospheric conditions. Effectene transfection reagent wasprepared as follows. 4 µl enhancer was added to 37 µl DNA condensation buffer (ECbuffer) and incubated at room temperature for 10 mins. 6 µl Effectene was thenadded and all reagents were vortexed before printing. Arrays were washed withMilliQ water to remove unbound DNA before cell culture.4-136