Mass transport effects on real-time nucleic acid microarrays

Abstract

Microarray analysis has become increasingly complex due to the growing size of arrays. In this work we explore the effects of diffusion and convective fluxes on the time of hybridization and sensing specificity in a single component system. A finite element software is used to simulate the diffusion of DNA through a microfluidic chamber to the sensing surface where hybridization of DNA is modeled using the corresponding kinetic equation. The differences between diffusion controlled and convection controlled mass transport are investigated as a function of concentration and hybridization time. Hybridization enhancement produced by microfluidics versus stationary diffusion is introduced as a useful metrics for quantitation of mass transport effects. MOTIVATIONSystemic approaches in genetic research, molecular diagnostics, and pharmacogenetics motivate the need for multi-target nucleic acid arrays with high sensitivity and selectivity. Arrays are also needed in the clinical application of genetic screening for the rapid identification of genetic disorders in the presence of multiple genotypes and/or mutations. 1, 2 Numerous statistical models have been created to evaluate the hybridization of nucleic acid targets using end-point analysis. 3, 4 Additionally, several authors have proposed mechanistic models of single component hybridization combining mass transport of target and chemical interactions on the surface of the array. [5][6][7][8][9] In this work we explore the effects of target species concentration and the time of hybridization on reaction rates in a single component systems with and without convection. A finite element method is used to simulate the diffusion and convection of DNA through a microfluidic chamber to the sensing surface where hybridization of DNA is modeled using the corresponding chemical reaction equation. The association rate constant for the 20-mer sequences is obtained from experimental data, for the perfectly matched target, and is used along with a thermodynamic model to determine dissociation rate constants. Virtual experiments were run for 60 minute time intervals for the concentration range of 1 pM -10 nM and wide range of flow rates. Additionally, experimental verification was completed using BioMicro System's mixing platform. MATHEMATICAL MODEL: SINGLE SPECIESNucleic acid arrays employ affinity interactions between complementary strands as the mechanism for sensing. Target which is delivered to the sensing surface from solution via diffusion and convection is interrogated by probes tethered to the sensing surface. The delivery of analyte to the surface is governed by Fick's Law.where C is the concentration of target in solution and D is the diffusion coefficient for the target, u is the convection flux. The affinity interaction at the sensing zone, in the case of a single species, can be expressed by

Theoretical limitations on sensing selectivity in nucleic acid microarrays

“…11,22 The mass transport model of both species is described by a system of independent Fick's equations.…”