title: Biochips for cell biology by combined dip-pen nanolithography and DNA-directed protein immobilization.
abstract: A general methodology for patterning of multiple protein ligands with lateral dimensions below those of single cells is described. It employs dip pen nanolithography (DPN) patterning of DNA oligonucleotides which are then used as capture strands for DNA-directed immobilization (DDI) of oligonucleotide-tagged proteins. This study reports the development and optimization of PEG-based liquid ink, used as carrier for the immobilization of alkylamino-labeled DNA oligomers on chemically activated glass surfaces. The resulting DNA arrays have typical spot sizes of 4-5 μm with a pitch of 12 μm micrometer. It is demonstrated that the arrays can be further functionalized with covalent DNA-streptavidin (DNA-STV) conjugates bearing ligands recognized by cells. To this end, biotinylated epidermal growth factor (EGF) is coupled to the DNA-STV conjugates, the resulting constructs are hybridized with the DNA arrays and the resulting surfaces used for the culturing of MCF-7 (human breast adenocarcinoma) cells. Owing to the lateral diffusion of transmembrane proteins in the cell's plasma membrane, specific recruitment and concentration of EGF receptor can be induced specifically at the sites where the ligands are bound on the solid substrate. This is a clear demonstration that this method is suitable for precise functional manipulations of subcellular areas within living cells.
authors.affiliation: Technische Universität Dortmund, Fakultät Chemie, Biologisch-Chemische Mikrostrukturtechnik, Otto Hahn Str. 6, 44227 Dortmund, Germany; Karlsruhe Institute of Technology (KIT), Institute for Biological Interfaces (IBG 1), Hermann-von-Helmholtz-Platz, D-76344 Eggenstein-Leopoldshafen, Germany; Scuola Superiore di Catania, Via Valdisavoia 9, 95123 Catania, Italy.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Arrabito
authors.forename: Stephanie
authors.initials: S
authors.lastname: Reisewitz
authors.forename: Leif
authors.initials: L
authors.lastname: Dehmelt
authors.forename: Philippe I
authors.initials: PI
authors.lastname: Bastiaens
authors.forename: Bruno
authors.initials: B
authors.lastname: Pignataro
authors.forename: Hendrik
authors.initials: H
authors.lastname: Schroeder
authors.forename: Christof M
authors.initials: CM
authors.lastname: Niemeyer
mesh_terms: D019921:Biotinylation; D003585:Cell Biology; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D004247:DNA / Q000737:chemistry*; D004815:Epidermal Growth Factor / Q000737:chemistry*; D005898:Glass / Q000737:chemistry; D049452:Green Fluorescent Proteins / Q000737:chemistry; D006801:Humans; D008024:Ligands; D061986:MCF-7 Cells; D008422:Materials Testing; D046208:Microchip Analytical Procedures; D036103:Nanotechnology; D020411:Oligonucleotide Array Sequence Analysis; D009841:Oligonucleotides / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry*; D013499:Surface Properties
chemical_list: D008024:Ligands; D009841:Oligonucleotides; D011506:Proteins; D049452:Green Fluorescent Proteins; D004815:Epidermal Growth Factor; D004247:DNA
keywords: DNA; cell biology; dip-pen nanolithography; microstructures; self-assembly