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Electric field pulse addressing of DNA, protein and cell-chips
Introduction
The objective of this research is to control the spatial localization and biochemical reactions of biological entities such as DNA, protein molecules, and cells. This work consists of: (i) the use of single voltage pulses to control the immobilization and hybridization of DNA; (ii) extension of the application of voltage pulses to control the kinetics antibody-antibody and antibody-antigen reactions; (iii) extension of the application of voltage pulses to achieve localized genetic uptake by cells through microelectroporation; (iv) integration of microfluidics in the biochips.
For more information on this topic, contact João Pedro Conde or Virginia Chu.
Main Results
The main result in this work up to now is the demonstration of control of the covalent immobilization and hybridization of DNA probes using ultra-short voltage pulses
Single, square voltage pulses in the microsecond time-scale result in selective 5´-end covalent bonding (immobilization) of thiolated single stranded (ss) DNA probes to a modified silicon dioxide flat surface and in specific hybridization of ssDNA targets to the immobilized probe. Immobilization and hybridization rates using microsecond voltage pulses at or below 1 V are at least 108 times faster than in the passive control reactions performed without electric field, and can be achieved with at least three differently functionalized thin film surfaces on plastic or glass substrates. The systematic study of the effect of DNA probe and target concentrations, of DNA probe and target length, and the application of asymmetric pulses on E-assisted DNA immobilization and hybridization showed that: (1) the rapidly rising edge of the pulse is most critical to the E-assisted processes, but the duration of the pulse is also important; (2) E-assisted immobilization and hybridization can be performed with micron-sized pixels, proving the potential for use on microelectronic length scales, and the applied voltage can be scaled down together with the electrode spacing to as low as 25 mV; and (3) longer DNA chains reduce the yield in the E-assisted immobilization and hybridization because the density of physisorbed single stranded DNA is reduced. The results show that the E-induced reactions can be used as a general method in DNA microarrays to produce high-density DNA chips (E-immobilization) and speed the microarray-based analysis (E-hybridization).
Recently, control of the hybridization of complementary DNA targets to covalently immobilized and electrostatically adsorbed DNA probes using single, square and sinusoidal voltage pulses in the microsecond time-scale has been achieved, resulting in specific hybridization of ssDNA targets to an probe electrostically immobilized to an APTES-functionalized surface.
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Schematic top-view of the test device (a) with mm-size metal electrodes. Schematic diagram of the passivation and chemically functionalized active layers (b). |
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Dependence of E-assisted DNA probe immobilization (a) and hybridization (b) on the duration of the single electric-field pulse (tp). |
Projects and Collaborations 2004 - present
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This work is done in collaboration with the Biological Engineering Research Group (BERG), of the Institute for Biotechnology and Bioengineering (IBB) (Miguel Prazeres, Gabriel Monteiro, Joaquim Sampaio Cabral).
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“Electronically-addressed biochips (eBioArray)” (POCTI/BIO/60487/2004). Partners: INESC-MN, BERG.
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"Development of microelectronically addressed DNA chips as generic platforms for genomic analysis" (POCTI/1999/BIO/34022). Partners: INESC-MN, BERG.