Electrical and Chemical Control of Surfaces for DNA Immobilization and Hybridization
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1093-CC04-23
Electrical and Chemical Control of Surfaces for DNA Immobilization and Hybridization R. Cabeça1,2, D. M.F. Prazeres2,3, V. Chu1, and J. P. Conde1,3 1 INESC-MN, Rua Alves Redol, 9, Lisbon, 1000-029, Portugal 2 IBB, IST, Av. Rovisco Pais, Lisbon, 1049-001, Portugal 3 Departament of Chemical and Biological Engineering, IST, Av. Rovisco Pais, Lisbon, 1049001, Portugal ABSTRACT We present the design of two biointerfaces on a SiO2 substrate for single stranded DNA (ssDNA) immobilization using either covalent grafting or electrostatic adsorption. The influence of the type of biointerface on the rate of diffusion-limited hybridization reaction with complementary ssDNA from a solution is studied. Patterning of the biointerface functionalization layers and the scaling down of the reaction volumes to μL range is demonstrated. The use of externally applied electric field pulses is shown to accelerate the hybridization reaction kinetics to the sub-ms time scale. INTRODUCTION As biological knowledge based on genomic studies increases, the use of DNA microarrays to identify gene expression in a massively parallel way is becoming indispensible. These devices are based on solid-phase hybridization of DNA targets from a solution to an ordered array of different DNA probes immobilized onto a chemically modified solid surface. Consequently, the design of the surface layer supporting the DNA probe, which includes the choice of the chemistry used for DNA immobilization [1], the control of the surface DNA probe density [2] and probe conformation, and the electrostatic interactions of the probe in the vicinity of the surface [3], is of major importance to improve the specificity of the hybridization reaction. Externally applied electric fields are extensively used for manipulation of DNA strands, e.g. in electrophoresis [4] and dielectrophoresis [5]. The complex behaviour exhibited by the highly charged DNA molecules when exposed to an electric field depends on the strand length [6], the ionic properties of the electrolyte solution, and the magnitude of the applied electric field [7]. Electric-field induced solid-phase hybridization on the sub-ms time range is shown to have yields comparable to those obtained for the equivalent passive reactions (without electric field). The influence of the shape (square vs. sinusoidal) and duration of an applied single voltage pulse on the kinetics of the hybridization reaction of ssDNA targets are presented for two types of biointerface layers of ssDNA probes immobilized by either covalent grafting or electrostatic adsorption to the surface. This technology can be integrated on microarray and lab-on-a-chip systems allowing on-chip addressing of DNA.
EXPERIMENT Surface functionalization For covalent immobilization of ssDNA probes (figure 1(a)), the SiO2 layer is submitted to a chemical functionalization protocol based on a silanization reaction with a 2% (v/v) solution of (3-aminopropyl)triethoxysilane, APTES (Fluka), prepared in acetone and a cross-linking reaction with a 1 mM solution o
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