Leo J. van IJzendoorn

One-step homogeneous magnetic nanoparticle immunoassay for biomarker detection directly in blood plasma.

Assay technologies capable of detecting low biomarker concentrations in complex biological samples are fundamental for biological research and for applications in medical diagnostics. In this paper we address the challenge to perform protein biomarker detection homogeneously in one single step, applying a minute amount of reagent directly into whole human blood plasma, avoiding any sample dilution, separation, amplification, or fluid manipulation steps. We describe a one-step homogeneous assay technology based on antibody-coated magnetic nanoparticles that are spiked in very small amount directly into blood plasma. Pulsed magnetic fields and a double-linker molecular architecture are used to generate high biomarker-induced binding and low nonspecific binding between the nanoparticles. We demonstrate dose-response curves for prostate specific antigen (PSA) measured in undiluted human blood plasma with a detection limit of 400-500 femtomol/L, in a total assay time of 14 min and an optically probed volume of only 1 nL. We explain the dose-response curves with a model based on discrete binding of biomarker molecules onto the nanoparticles, which allows us to extract reaction parameters for the binding of biomarker molecules onto the nanoparticles and for the biomarker-induced binding between nanoparticles. The demonstrated analytical performance and understanding of the nanoparticle assay technology render it of interest for a wide range of applications in quantitative biology and medical diagnostics.

Reaction-diffusion model for the preparation of polymer gratings by patterned ultraviolet illumination

A model is developed to describe the migration mechanism of monomers during the lithographic preparation of polymer gratings by ultraviolet polymerization. The model is based on the Flory–Huggins theory: a thermodynamic theory that deals with monomer/polymer solutions. During the photoinduced polymerization process, monomer migration is assumed to be driven by a gradient in the chemical potential rather than the concentration. If the chemical potential is used as the driving force, monomer migration is not only driven by a difference in concentration, or volume fraction, but also by other entropic effects such as monomer size and the degree of crosslinking of the polymer network, which is related to the ability of a polymer to swell. Interaction of the monomers with each other or the polymer is an additional energetic term in the chemical potential. The theoretical background of the model is explained and results of simulations are compared with those of nuclear microprobe measurements. A nuclear micropro...

Confined Brownian motion of individual magnetic nanoparticles on a chip: Characterization of magnetic susceptibility.

An increasing number of biomedical applications requires detailed knowledge of the magnetic susceptibility of individual particles. With conventional techniques it is very difficult to analyze individual particles smaller than 1μm. The authors demonstrate how the susceptibility of individual nanoparticles can be determined in an efficient way by optically analyzing the confined Brownian motion of a nanoparticle trapped in a known magnetic potential well on a chip. A setup is introduced that has a controllable two-dimensional magnetic potential well, which is defined by an integrated microscopic current wire. Susceptibility measurements have been performed on 150–450nm superparamagnetic beads. They found differences in bead susceptibility of an order of magnitude and differences in volumetric susceptibility of more than a factor of 2.

Sensitivities of InGaAsP photonic crystal membrane nanocavities to hole refractive index

The sensitivities of resonant wavelengths of photonic crystal (PhC) membrane nanocavities with embedded InAs quantum dots to the ambient refractive index are reported for use in (bio) chemical sensing. The resonances for the different modes of several point-defect type cavities are obtained by photoluminescence measurements. Systematic trends of the variation of sensitivity with increase of the overlap of the modes with the PhC holes are observed for varying cavity type as well as for a given mode within a cavity type. A maximum sensitivity of approximately 300 nm/RIU (refractive index unit) is observed, corresponding to approximately 25% mode overlap with the holes and complete infiltration with the aqueous solution.

Simulations with a dynamic reaction-diffusion model of the polymer grating preparation by patterned ultraviolet illumination

Simulations of volume fraction profiles formed during the lithographic preparation of polymer gratings are made with a reaction/diffusion model, based on the Flory–Huggins theory. Monomer migration is driven by a gradient in the chemical potential rather than a gradient in the concentration. If the chemical potential is used as the driving force, monomer migration is not only driven by a difference in concentration, or volume fraction, but also by other entropic effects: the differences in monomer length and the degree of crosslinking of a polymer network. The monomer volume fractions are simulated as a function of position for different ultraviolet intensities and various grating pitches. Profound edges of the monomer volume fractions caused by the fact that the reaction rate is high compared to the diffusion rate are both measured and simulated. An excellent agreement with nuclear microprobe measurements on the polymer gratings is obtained.

Quantification of protein-ligand dissociation kinetics in heterogeneous affinity assays

Biochemical affinity assays inherently involve interactions of heterogeneous nature. We report a methodology to discriminate between and accurately characterize specific and nonspecific interactions in force-induced dissociation assays. Ligand-coupled superparamagnetic particles are incubated on surfaces coated with a mixture of specific receptors and nonspecifically interacting proteins. Consequently, a mixed population of surface bound particles is formed with different binding natures. Magnetic field gradients are used to apply translational forces on the bound particles. Using a multicomponent dissociation analysis, we are able to make a distinction between weak nonspecific interactions, strong nonspecific interactions, and specific interactions. We validate the model by comprehensive experiments in which the biochemical components and applied forces are varied. The low-force data yield reliable values for the spontaneous dissociation rates of single-molecule specific bonds, and at high forces, the bond barriers are modified by the applied force. The results generate a new perspective for applications of magnetic force affinity assays in studies of heterogeneous molecular biorecognition.

Analysis of individual magnetic particle motion near a chip surface

We describe an analysis of the dynamics of individual superparamagnetic micro- and nanoparticles in order to quantify their magnetic properties and mobility near a chip surface. The particles are attracted to the chip surface by integrated microscopic current wires. We show that it is possible to accurately analyze particles with a diameter of about 1 μm by the magnetophoretic movement between current wires because of the very high field gradients. This reveals distinct differences in volume susceptibilities of particles with the same outer diameter. Smaller particles are characterized using the technique of confined Brownian motion analysis. By capturing 300 nm particles on a current wire with surface barriers or a focused shape, the magnetization of the particles can be measured with an accuracy better than 10%.

Probing the force-induced dissociation of aptamer-protein complexes.

Aptamers are emerging as powerful synthetic bioreceptors for fundamental research, diagnostics, and therapeutics. For further advances, it is important to gain a better understanding of how aptamers interact with their targets. In this work, we have used magnetic force-induced dissociation experiments to study the dissociation process of two different aptamer-protein complexes, namely for hIgE and Ara h 1. The measurements show that both complexes exhibit dissociation with two distinct regimes: the dissociation rate depends weakly on the applied force at high forces but depends stronger on force at low forces. We attribute these observations to the existence of at least one intermediate state and at least two energy barriers in the aptamer-protein interaction. The measured spontaneous dissociation rate constants were validated with SPR using both Biacore and fiber optic technology. This work demonstrates the potential of the magnetic force-induced dissociation approach for an in-depth study of the dissociation kinetics of aptamer-protein bonds, which is not possible with SPR technologies. The results will help in the development and expansion of aptamers as bioaffinity probes.

Roughness and ordering at the interface of oxidized polystyrene and water

For the first time, atomistically detailed molecular dynamics calculations revealed molecular ordering of the water-oxidized atactic polystyrene (aPS) interface. Both ordering of the water molecules and the phenyl rings occur. In addition, the natural roughness of the surface has been simulated and compared to experimental values. The composition of the simulated aPS films is based on spin-coated aPS films that have been oxidized and characterized experimentally. The aPS surfaces are oxidized with ultraviolet-ozone radiation and have been characterized by XPS, AFM, and water contact angle measurements. XPS measurements show that the oxygen content in the sample increases rapidly with exposure and reaches saturation near 24 at. % of oxygen. The molecular dynamics simulations show smoothening of an hydrophobic aPS surface upon transition from vacuum to water. The smoothening decreases with increasing hydrophilicity. The calculations reveal ordering of oxidized phenyl rings for aPS surfaces in water. The order increases with increasing hydrophilicity. Additionally, we investigated the water structure near the aPS-water interface as a function of the surface hydrophilicity. With increasing hydrophilicity, the density of water at the aPS-water interface increases. The water density profile is steeper in the presence of hydrophobic aPS. The water shows an ordered layer near both the hydrophobic and hydrophilic surfaces; the position of this layer shifts toward the interface with increasing hydrophilicity.

Mobility and height detection of particle labels in an optical evanescent wave biosensor with single-label resolution

Particle labels are used in biosensors to detect the presence and concentration of analyte molecules. In this paper we demonstrate an optical technique to measure the mobility and height of bound particle labels on a biosensor surface with single-label resolution. The technique is based on the detection of the particle-induced light scattering in an optical evanescent field. We show that the thermal particle motion in the optical evanescent field leads to intensity fluctuations that can accurately be detected. The technique is demonstrated using 290 bp (99 nm) DNA as an analyte and using polystyrene particles and magnetic particles with diameters between 500 and 1000 nm as labels. The particle intensity histograms show that quantitative height measurements are obtained for particles with uniform optical properties, and the intensity versus position plots reflect the analyte–antibody orientation and the analyte flexibility. The novel optical detection technique will lead to biosensors with very high sensitivity and specificity.