title: Microscale mesoarrays created by dip-pen nanolithography for screening of protein-protein interactions.
abstract: Using microarrays to probe protein-protein interactions is becoming increasingly attractive due to their compatibility with highly sensitive detection techniques, selectivity of interaction, robustness and capacity for examining multiple proteins simultaneously. The major drawback to using this approach is the relatively large volumes and high concentrations necessary. Reducing the protein array spot size should allow for smaller volumes and lower concentrations to be used as well as opening the way for combination with more sensitive detection technologies. Dip-Pen Nanolithography (DPN) is a recently developed technique for structure creation on the nano to microscale with the capacity to create biological architectures. Here we describe the creation of miniaturised microarrays, 'mesoarrays', using DPN with protein spots 400× smaller by area compared to conventional microarrays. The mesoarrays were then used to probe the ERK2-KSR binding event of the Ras/Raf/MEK/ERK signalling pathway at a physical scale below that previously reported. Whilst the overall assay efficiency was determined to be low, the mesoarrays could detect KSR binding to ERK2 repeatedly and with low non-specific binding. This study serves as a first step towards an approach that can be used for analysis of proteins at a concentration level comparable to that found in the cellular environment.
authors: Centre for Molecular Nanometrology, Department of Pure and Applied Chemistry, University of Strathclyde, Glasgow, G1 1XL, UK.
authors: David G
authors: DG
authors: Thompson
authors: Ekaterina O
authors: EO
authors: McKenna
authors: Andrew
authors: A
authors: Pitt
authors: Duncan
authors: D
authors: Graham
mesh_terms: D000818:Animals; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D036103:Nanotechnology; D040081:Protein Array Analysis / Q000295:instrumentation*; D011485:Protein Binding; D025941:Protein Interaction Mapping / Q000295:instrumentation*; D011494:Protein Kinases / Q000378:metabolism*; D012680:Sensitivity and Specificity
chemical_list: D011494:Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1