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Kris Pappaert

Publications and source records attributed to Kris Pappaert.

9 recordsLinked to original sources
PubMed2006

A dimensionless number analysis of the hybridization process in diffusion- and convection-driven DNA microarray systems.

The present theoretical analysis aims at providing a general understanding of the combined effect the many different process variables have on the hybridization rate in diffusion- and convection-driven DNA microarray systems. It is shown that all process variables can be grouped into only four different dimensionless numbers (the Damkohler number Da, the dimensionless association constant kappa(A), the dimensionless initial concentration C'(0) and a geometrical ratio alpha). These four numbers have a straightforward physical meaning and only contain easily measurable parameters. Reducing the solution space from 7D to 4D, the dimensionless number representation greatly facilitates the insight in the conditions leading to the occurrence of diffusion-limited hybridization rates in both diffusion- and convection-driven DNA microarray systems. This in turn simplifies their design and the interpretation of the experimental results that are obtained with these systems.

Diffusion↗
PubMed2006

Diffusion limitation: a possible source for the occurrence of doughnut patterns on DNA microarrays.

Doughnut shaped hybridization patterns on DNA microarrays are mainly allocated to spotting or drying artifacts. The present study reports on results obtained from four different approaches that when combined generate a better view on the occurrence of these patterns. This study points out that doughnuts are not only formed during the spotting and drying process, but the hybridization process itself can be considered as an important cause. A combination of computer simulations, theoretical, optical, and experimental techniques shows how ring-shaped hybridization patterns occur when diffusion-limited conditions are present during the hybridization process. The theoretical assumptions as well as the simulations indicate that, for the basic geometry of a microarray hybridization experiment, a large amount of binding molecules reach the spot from the sides (and not from above the spot), leading to a preferential binding on the rims of the spot. These patterns seem to occur especially during hybridization with short oligonucleotides that have a very high binding probability and fast hybridization kinetics. Longer target DNA molecules lead to a more evenly distributed intensity signal. Furthermore, the diffusion-limited conditions also lead to pronounced hybridization intensity patterns on the scale of a whole spot block, where larger intensities are obtained on the edges of the block compared with the spots laying in the center of the block.

Animals↗
PubMed2005

Measurements of diffusion coefficients in 1-D micro- and nanochannels using shear-driven flows.

The present paper describes a method for measuring the molecular diffusion coefficient of fluorescent molecules in microfluidic systems. The proposed static shear-driven flow method allows one to perform diffusion measurements in a fast and accurate manner. The method also allows one to work in very thin (i.e. submicron) channels, hence allowing the investigation of diffusion in highly confined spaces. In the deepest investigated channels, the obtained results were comparable to the existing literature values, but when the channel size dropped below the micrometer range, a significant decrease (more than 30%) in molecular diffusivity was observed. The reduction of the diffusivity was most significant for the largest considered molecules (ssDNA oligomers with a size ranging between 25 to 100 bases), but the decrease was also observed for smaller tracer molecules (FITC). This decrease can be attributed to the interactions of the analyte molecules with the channel walls, which can no longer be neglected when the depth of the channel reaches a critical value. The change in diffusivity seems to become more explicit as the molecular weight of the analytes increases.

DNA, Single-Stranded↗
PubMed2005

DNA microarray enhancement using a continuously and discontinuously rotating microchamber.

It is demonstrated that the most efficient way to enhance DNA microarray analysis consists of a maximal reduction of the total device volume (to keep the concentration of the available DNA as high as possible), combined with the creation of a strong lateral convective transport of the sample. In the present study, DNA microarray hybridizations are performed in a set of rotating, circular microchambers covering exactly the spotted area of the microarray and with a depth varying between 70 and 1.6 microm. Rotating the microchamber substrate while keeping the microarray stationary, the rotating microchamber bottom wall literally drags the sample past the microarray spots with a velocity which is independent of the fluid layer thickness. Interestingly, it was found that transporting the sample in a discontinuous mode (with stop periods of several minutes) not only yields a more stable and reproducible operation, it also yields significantly larger hybridization intensities (typically a factor of 2-3 larger) than a continuous rotation. This seems to be due to the fact that the velocity field disturbs the binding process at the binding site level. Working under limiting DNA sample mass conditions, the system yielded in a short, 30-min experiment already a 5-fold increase of the hybridization intensity, as compared to a conventional microscope slide/coverslip system operated overnight under diffusion-driven conditions. Compared to a commercial pump-around hybridization system, the gain was even more impressive, precisely due to the fact that the pump-around system requires larger volumes, which with a fixed amount of available genetic material leads to the application of more diluted samples.

DNA, Complementary↗
PubMed2005

Comparison of a pump-around, a diffusion-driven, and a shear-driven system for the hybridization of mouse lung and testis total RNA on microarrays.

In the present study, we demonstrate the benefits of a shear-driven rotating microchamber system for the enhancement of microarray hybridizations, by comparing the system with two commonly used hybridization techniques: purely diffusion-driven hybridization under coverslip and hybridization using a fully automated hybridization station, in which the sample is pumped in an oscillating manner. Starting from the same amount of DNA for the three different methods, a series of hybridization experiments using mouse lung and testis DNA is presented to demonstrate these benefits. The gain observed using the rotating microchamber is large: both in terms of analysis speed (up to tenfold increase) and in final spot intensity (up to sixfold increase). The gain is due to the combined effect of the hybridization chamber miniaturization (leading to a sample concentration increase if comparing iso-mass conditions) and the transport enhancement originating from the rotational shear-driven flow induced by the rotation of the chamber bottom wall.

Animals↗
PubMed2004

High-velocity transport of nanoparticles through 1-D nanochannels at very large particle to channel diameter ratios.

We explore the possibility of generating high-velocity flows of nanoparticles through flat-rectangular nanochannels, which are only 50% deeper than the diameter of the particles. Using the shear-driven flow principle, 200-nm particles can, for example, be transported through a 300-nm-deep channel at velocities up to 35 mm/s (upper limit of our current setup). Working under high-pH conditions, the velocity of the carboxylated nanoparticles still respects the small-molecule velocity law, despite the high degree of confinement to which the particles are subjected. The high degree of confinement is also found to lead to a reduced band broadening. When injecting sharply delimited particle plugs, the plate heights observed for the flow of 0.2-microm particles through a 0.3-microm channel (with plate heights of the order of 1-2 microm) are, for example, approximately 1 order of magnitude smaller than for the flow of 1.0-microm particles through a 1.4-microm channel. It is also found that the band broadening is, within its statistical variation, independent of the fluid velocity over a large range of particle velocities (5-35 mm/s). The flow method distinguishes itself from pressure-driven field-flow fractionation and hydrodynamic chromatography in that the mean particle velocity is independent of the particle size over the entire range of possible particle to channel diameter ratios.

Journal Article↗
PubMed2004

A novel microstep device for the size separation of cells.

We report on a series of preliminary experiments investigating the applicability of a novel method for the size separation of nano- and microsized particles and cells. The working principle is based on the application of a shear-driven flow through stepwise tapered micro- or nanochannels. Size separations of mixtures of 0.5 and 1.0 microm carboxylated polystyrene beads as well as of binary mixtures of Staphylococcus aureus and Saccharomyces cerevisiae cells and of S. cerevisiae and Escherichia coli cells are demonstrated.

Escherichia coli↗
PubMed2004

Exploiting the benefits of miniaturization for the enhancement of DNA microarrays.

The present study demonstrates that the best way to enhance DNA microarray assays, both in terms of analysis speed and in final spot intensity, is to dissolve the available molar amount of sample in the smallest possible buffer volume and to subsequently convect this solution continuously across the surface of the array. The presently proposed shear-driven flow system is pre-eminently suited for this task, as it allows to induce strongly enhanced lateral transport rates, independently of the degree of miniaturization of the hybridization chamber. This transport enhancement method, however, only increases the hybridization rate and not the final spot intensity, as neither can any of the other transport enhancement methods already proposed in literature. A series of experiments with synthetic single-stranded (ssDNA) samples and an accompanying mass balance analysis are presented to demonstrate these points.

DNA, Single-Stranded↗