Tuned graft copolymers as controlled coatings for DNA microarrays.
DNA microarrays have become a powerful tool for expression profiling and other genomics applications. A critical factor for their sensitivity is the interfacial coating between the chip substrate and the bound DNA. Such a coating has to embrace the divergent requirements of tightly binding the capture probe DNA during the spotting process and of minimizing the nonspecific binding of target DNA during the hybridization assay. To fulfill these conditions, most coatings require a passivation step. Here we demonstrate how the chain density of a graft copolymer with a polycationic backbone, poly(l-lysine)-graft-poly(ethylene glycol), can be tuned such that the binding capacity during capture probe deposition is maximized while the nonspecific binding during hybridization assays is kept to a minimum, thus alleviating the requirement for a separate passivation procedure. Evidence for the superior performance of such coatings in terms of signal-to-noise ratio and spot quality is presented using an evanescent field-based fluorescent sensing technique (the ZeptoREADER). The surface architecture is further characterized using optical waveguide lightmode spectroscopy and time-of-flight secondary ion mass spectrometry. Finally, in a model assay, we demonstrate that expression changes can be detected from 1 microg of total mRNA sample material with a limit of detectable differential expression of +/-1.5.
De Paul
Susan M
SM
Laboratory for Surface Science and Technology, Department of Materials, Swiss Federal Institute of Technology, ETH-Zürich, Wolfgang-Pauli-Strasse 10, CH-8093 Zürich, Switzerland.
Falconnet
Didier
D
Pasche
Stéphanie
S
Textor
Marcus
M
Abel
Andreas P
AP
Kauffmann
Ekkehard
E
Liedtke
Roman
R
Ehrat
Markus
M
D004247:DNA / Q000737:chemistry*; D013058:Mass Spectrometry; D046208:Microchip Analytical Procedures; D015394:Molecular Structure; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D009841:Oligonucleotides / Q000737:chemistry; D009994:Osmolar Concentration; D011092:Polyethylene Glycols / Q000737:chemistry*; D011107:Polylysine / Q000737:chemistry; D055672:Static Electricity; D013379:Substrate Specificity
D009841:Oligonucleotides; D011107:Polylysine; D011092:Polyethylene Glycols; D004247:DNA