Novel polymeric coatings with tailored hydrophobicity to control spot size and morphology in DNA microarray

Abstract

The fabrication of high density microarrays requires a precise control of surface physical and chemical properties. In fact, spot morphology has a deep impact on signal level and might affect seriously the reliability and reproducibility of the assay. As a consequence, a proper biomolecule deposition and immobilization increases the accuracy of data analysis. Here we propose a new polymeric coating with intermediate hydrophilic/hydrophobic characteristics able to control the spreading of spots onto the surface as well as their size, thus enabling the reduction of the pitch between spots. The new coating exemplifies the versatility of a copolymer system based on the simultaneous presence of N,N-Dimethylacrylamide (DMA), 3-(trimethoxylsilyl)propyl methacrylate (MAPS) and fluorinated monomers, which impart to the surface a more hydrophobic behavior, reducing the drops spreading and merging. The controlled hydrophobicity allows to control also the size of the spots, thus the possibility of reducing their pitch to 140μm and to obtain arrays with smaller dimentions, a very useful characteristics in those applications which imply the use of reduced areas, for example in miniaturized biosensors or microfluidic devices. The obtained copolymer is easily adsorbed onto glass or silicon oxide from a diluted aqueous solution of the polymer and the result is a microarray surface with intermediate hydrophobic/hydrophilic properties (average water contact angle is 50° and surface energy values are reduced from 114mN/m to 50mN/m), which allows decrease of the spot distance without merging of the droplets. To further improve the spot size and morphology, we have also investigated the use of several detergents added to the spotting buffer: in particular, the use of sucrose monolaurate, together with the hydrophobic surface, permitted the fabrication of surfaces for dense DNA microarray with very high fluorescence signals and low background noise. In fact, as an example of application in DNA microarray, the fluorinated coating has been used for the genotyping of KRAS G12D mutation, a common variant in the KRAS gene implicated in the colorectal cancer.