Materials Screening for Sol-Gel-Derived High-Density Multi-Kinase Microarrays

摘要

Protein microarrays based on pin-printing of sol—gel-entrapped biomolecules have emerged as a potential tool to accelerate drug screening and discovery. However, while materials have recently been identified that are suitable for printing of high-density sol—gel-based microarrays, the ability to print arrays of delicate proteins such as kinases, and to assay their activity and inhibition on-array, has yet to be demonstrated. In this study, we have performed a criteria-based directed screen of sol—gel-based materials to identify compositions that are suitable for the fabrication of high-density, multikinase microarrays. Printable formulations were assessed for their compatibility with a fluorescent, phosphospecific dye used as an end-point indicator for on-array kinase assays, including an assessment of the effects of spot size (l00 μm vs 400 μm) and slide surface chemistry on signal reproducibility. The combinations of materials, surfaces, and spot sizes that were found to be compatible with reproducible signal generation were evaluated for their ability to retain the activity of a range of kinases, which were co-entrapped with their respective substrates into the optimal sol—gel materials to produce microarrays. Ultimately, two material/surface combinations, from potentially thousands, were identified, one of which was used to produce a robust, highly active kinase microarray that could be used for qualitative screening as well as quantitative inhibition assays.