Tailored magnetic nanoparticles for direct and sensitive detection of biomolecules in biological samples.

We developed nanoparticles with tailored magnetic properties for direct and sensitive detection of biomolecules in biological samples in a single step. Thermally blocked nanoparticles obtained by thermal hydrolysis, functionalized with specific ligands, are mixed with sample solutions, and the variation of the magnetic relaxation due to surface binding is used to detect the presence of biomolecules. The binding significantly increases the hydrodynamic volume of nanoparticles, thus changing their Brownian relaxation frequency which is measured by a specifically developed AC susceptometer. The system was tested for the presence of Brucella antibodies, a dangerous pathogen causing brucellosis with severe effects both on humans and animals, in serum samples from infected cows and the surface of the nanoparticles was functionalized with lipopolysaccharides (LPS) from Brucella abortus. The hydrodynamic volume of LPS-functionalized particles increased by 25-35% as a result of the binding of the antibodies, measured by changes in the susceptibility in an alternating magnetic field. The method has shown high sensitivity, with detection limit of 0.05 microg x mL(-1) of antibody in the biological samples without any pretreatment. This magnetic-based assay is very sensitive, cost-efficient, and versatile, giving a direct indication whether the animal is infected or not, making it suitable for point-of-care applications. The functionalization of tailored magnetic nanoparticles can be modified to suit numerous homogeneous assays for a wide range of applications.

QCM-D analysis of the performance of blocking agents on gold and polystyrene surfaces.

With today's developments of biosensors and medical implants comes the need for efficient reduction of nonspecific binding. We report on a comparison of the ability of traditionally used blocking agents and poly(ethylene glycol) (PEG) derivatives to prevent protein adsorption on both gold and polystyrene surfaces. The adsorption kinetics of blocking molecules and proteins was monitored gravimetrically using quartz crystal microbalance with dissipation (QCM-D). The resistance to nonspecific adsorption was evaluated on gold and polystyrene surfaces coated with bovine serum albumin (BSA) or casein, gold coated with three different 6-11 ethylene glycol (EG) long hydroxyl- or methoxy-terminated PEG-thiolates and polystyrene blocked with a PLL-g-PEG or three different 12 EG long benzyl-PEG-derivatives. The prevention of protein adsorption on the coated surfaces was evaluated by monitoring the mass uptake at the addition of both pure prostate specific antigen (PSA) and seminal plasma. We demonstrate that on pure gold the PEG-thiols are superior to the other blocking molecules tested, with the end group and length of the PEG-thiols used being of minor importance. On polystyrene surfaces blocking with PLL-g-PEG, BSA and casein gave the best results. These results have an impact on further development of an optimized immunoassay protocol.

Brownian motion of aggregating nanoparticles studied by photon correlation spectroscopy and measurements of dynamic magnetic properties.

We have investigated colloidal stability of magnetic nanoparticle suspensions in different buffer systems and NaCl concentrations commonly used for biological applications. We have also investigated how conjugation of proteins to magnetic nanoparticles affects colloidal stability. Two different techniques, giving complementary information on the state of the particle system studied, have been used and compared. We have monitored the rotational Brownian motion of particles using measurements of dynamic magnetic susceptibility in the frequency domain. The results were processed using an algorithm that enables us to quantify changes of particle size distribution for particle suspensions subjected to various buffer conditions. The measurements were compared to results obtained for the translational Brownian motion of the same nanoparticles using photon correlation spectroscopy (PCS). We demonstrate that the complementarity of the two techniques enables more precise characterization of particles in suspension, particularly for suspensions of particles with a wide distribution in size and shape, or systems that are close to the onset of agglomeration.