Magnetoresistive sensors for measurements of DNA hybridization kinetics - effect of TINA modifications.
We present the use of magnetoresistive sensors integrated in a microfluidic system for real-time studies of the hybridization kinetics of DNA labeled with magnetic nanoparticles to an array of surface-tethered probes. The nanoparticles were magnetized by the magnetic field from the sensor current. A local negative reference ensured that only the specific binding signal was measured. Analysis of the real-time hybridization using a two-compartment model yielded both the association and dissociation constants kon, and koff. The effect of probe modifications with ortho-Twisted Intercalating Nucleic Acid (TINA) was studied. Such modifications have been demonstrated to increase the melting temperature of DNA hybrids in solution and are also relevant for surface-based DNA sensing. Kinetic data for DNA probes with no TINA modification or with TINA modifications at the 5' end (1 × TINA) or at both the 5' and 3' ends (2 × TINA) were compared. TINA modifications were found to provide a relative decrease of koffby a factor of 6-20 at temperatures from 57.5 °C to 60 °C. The values of konwere generally in the range between 0.5-2 × 105M-1s-1and showed lower values for the unmodified probe than for the TINA modified probes. The observations correlated well with measured melting temperatures of the DNA hybrids.
Department of Micro- and Nanotechnology, Technical University of Denmark, DTU Nanotech, Building 345B, DK-2800 Kongens Lyngby, Denmark.
G
G
Rizzi
Department of Micro- and Nanotechnology, Technical University of Denmark, DTU Nanotech, Building 345B, DK-2800 Kongens Lyngby, Denmark.
M
M
Dufva
Department of Micro- and Nanotechnology, Technical University of Denmark, DTU Nanotech, Building 345B, DK-2800 Kongens Lyngby, Denmark.
M F
MF
Hansen
D015374:Biosensing Techniques / Q000295:instrumentation*; D004247:DNA / Q000032:analysis* / Q000737:chemistry*; D006801:Humans; D007364:Intercalating Agents / Q000032:analysis* / Q000737:chemistry*; D007700:Kinetics; D008280:Magnetics / Q000295:instrumentation*; D009693:Nucleic Acid Hybridization; D044366:Transition Temperature
D007364:Intercalating Agents; D004247:DNA