Patterned and switchable surfaces for biomaterial applications

Chapter 2 – Spatially controlled electro-stimulated DNA adsorption and desorption for biodeviceapplications2.3.4. Formation of a transfected cell microarrayUtilising the ALAPP/PEG surfaces, patterned by laser ablation, for TCMapplications was of interest. After surface patterning pEGFP-N1 plasmid withEffectene transfection reagent was spotted onto the ALAPP regions using apiezoelectric non-contact printer. The surface was then seeded with HEK 293 cellsand allowed to incubate for 24 hr. A fluorescence microscopy image of the formedcell array is shown in Figure 2.9. Here the cells were stained with Hoechst dye,which stains the nucleus of all cells present. Cells attaching to different ALAPPregions are clearly separated from each other. The absence of a lawn of cells couldbe enormously beneficial for detecting subtle, subcellular phenotypic changes withintransfected cells, as there would be no need to differentiate between transfected andnon-transfected cells, provided that near 100% of cells within each spot aretransfected. Furthermore, as there is a migration barrier there is no way thattransfected cells or adsorbed DNA can migrate from one cell cluster to another. Thiscould allow for cell colonies to be positioned closer together allowing for thecreation of high-density arrays.The inset of Figure 2.9 shows an overlayed image of the fluorescence due to theHoechst dye and also GFP fluorescence due to the expression of the pEGFP-N1plasmid from a single cell cluster. As can be seen from the inset, some cells appeargreen, whilst others remain only blue, showing that 100% of cells were nottransfected. Typically, a transfection efficiency of 20% was achieved. Furtherimprovement to this transfection efficiency is possible by application of a voltageafter cell attachment and by optimising the amount of DNA and transfection reagent2-94