| Sensor and method for measuring the areal density of magnetic nanoparticles on a micro-array -> Monitor Keywords |
|
|
  Sensor and method for measuring the areal density of magnetic nanoparticles on a micro-arrayUSPTO Application #: 20060128035Title: Sensor and method for measuring the areal density of magnetic nanoparticles on a micro-array Abstract: A magnetoresistive sensor device for measuring an areal density of magnetic nanoparticles on a micro-array, the magnetic nanoparticles (15) being directly or indirectly coupled to the target sample (11), is described. The magnetoresistive sensor device comprises a substrate (3) having attached thereto binding sites (9) able to selectively bind target sample (11), and a magnetoresistive sensor for detecting the magnetic field of the nanoparticles (15) coupled to the target sample. The magnetoresistive sensor comprises a plurality of magnetoresistive sensing elements (17, 19), the width and length dimensions of which are at least a factor 10 or more, preferably a factor 100 or more larger than the diameter of the nanoparticles (15). A corresponding method is described as well. The present invention relates to a method and a device for magnetic detection of binding of biological molecules on a biochip. (end of abstract) Agent: Philips Intellectual Property & Standards - Briarcliff Manor, NY, US Inventors: Reinder Coehoorn, Menno Willem Jose Prins USPTO Applicaton #: 20060128035 - Class: 436524000 (USPTO) Related Patent Categories: Chemistry: Analytical And Immunological Testing, Involving An Insoluble Carrier For Immobilizing Immunochemicals, Carrier Is Inorganic The Patent Description & Claims data below is from USPTO Patent Application 20060128035. Brief Patent Description - Full Patent Description - Patent Application Claims
[0001] The present invention relates to a method and apparatus for sensing of randomly positioned nanometer-scale magnetic particles. In particular it relates to magnetic detection apparatus and a method for binding of biological molecules on a micro-array or biochip. [0002] The introduction of micro-arrays or biochips is revolutionising the analysis of DNA (desoxyribonucleic acid), RNA (ribonucleic acid) and proteins. Applications are e.g. human genotyping (e.g. in hospitals or by individual doctors or nurses), bacteriological screening, biological and pharmacological research. [0003] Biochips, also called biosensor chips, biological microchips, gene-chips or DNA chips, consist in their simplest form of a substrate on which a large number of different probe molecules are attached, on well defined regions on the chip, to which molecules or molecule fragments that are to be analysed can bind if they are perfectly matched. For example, a fragment of a DNA molecule binds to one unique complementary DNA (c-DNA) molecular fragment. The occurrence of a binding reaction can be detected, e.g. by using fluorescent markers that are coupled to the molecules to be analysed. This provides the ability to analyse small amounts of a large number of different molecules or molecular fragments in parallel, in a short time. One biochip can hold assays for 1000 or more different molecular fragments. It is expected that the usefulness of information that can become available from the use of biochips will increase rapidly during the coming decade, as a result of projects such as the Human Genome Project, and follow-up studies on the functions of genes and proteins. [0004] One method for electronically detecting binding of sample molecules to probe molecules has been demonstrated by Clinical Micro Sensors (CMS), a subsidiary of Motorola, and is described in D. H. Farkas, "Bioelectric detection of DNA and the automation of molecular diagnostics", The Journal of the Association for Laboratory Automation, volume 4, number 5 (1999), pp. 20-24. They have developed a "bioelectric DNA detection chip". The principle requires the use of ferrocene label molecules, which are sources or sinks of electrons. Capture probes are attached to gold-coated electrodes on the biochip. Capture probes are single strands of DNA complementary to a unique region of the target DNA or RNA sequence. When a sample containing target DNA is introduced into the cartridge, specific capture probes on an electrode surface encounter complementary DNA from the sample. Then binding, or hybridisation, occurs. The system also contains DNA sequences, called signaling probes, with proprietary electronic labels attached to them. These signaling probes also bind to the target DNA sequence. Binding of the target sequence to both the capture probe and the signaling probe connects the electronic labels to the surface. Binding of a molecular fragment is detected by the occurrence of an AC current through an electrode on which the molecules are bound, when a slight AC voltage is applied between the electrode and the solution above the chip, because the labels release electrons, producing a characteristic signal that can be detected through the electrode. This indicates the presence of the target DNA. Within this concept, the signal is proportional to the absolute number of binding reactions that have taken place. The number of electrons that flow, per cycle, and per bound DNA/c-DNA pair, is very small (a few, or a few tens). The above-mentioned paper mentions that in practice currents are in the pA to .mu.A range, unfortunately without specifying the electrode area or the absolute number of bound pairs (presumably very large numbers). Proprietary signal processing technology is used to identify and quantify the target DNA sequence. [0005] A second principle is a Bead Array Counter (BARC) biochip, as described in D. R. Baselt, "A biosensor based on magnetoresistance technology", Biosensors & Bioelectronics 13, 731-739 (1998); in R. L. Edelstein et al., "The BARC biosensor applied to the detection of biological warfare agents", Biosensors & Bioelectronics 14, 805 (2000); and in M. M. Miller et al., "A DNA array sensor utilizing magnetic microbeads and magnetoelectronic detection", Journal of Magnetism and Magnetic Materials 225 (2001), pp. 138-144. [0006] Using magnetoresistive materials, a rugged, single-component, micro-fabricated detector is produced, that will simultaneously monitor hundreds, thousands or even millions of experiments. As shown in FIG. 1, the detector 100 has an array of many micron-sized magnetoresistive sensors 101. For clarity, only two probe sites, each with one GMR sensor, are shown. These sensors 101 are located at a very small depth (=thickness of the silicon nitride Si.sub.3N.sub.4 passivation layer 102 combined with that of a relatively thin gold layer 103) below the surface of the substrate to which two probe DNA 104 is attached. Biotinylated sample DNA 105 binds to the probe DNA 104, if the nucleotide sequences the pairs formed are complementary. To the solution free-floating magnetic streptavidin coated micro-beads 106 are added, after probe-sample DNA hybridization has taken place. The beads bind to the sample DNA that has hybridized with probe DNA by the formation of a streptavidin-biotinyl bond. So if sample DNA 105 having sequences complementary to both probes 104 is present, the sample DNA 105 will attach the beads 106 to the sensors 101. The beads 106 used have a diameter of the order of 1 .mu.m. In the beads 106, nanometer scale magnetic particles are present (not represented in the drawing), which are superparamagnetic due to their small size. Those nanometer-sized particles are typically of iron oxide, and are dispersed in, layered onto, or coated with a polymer or silica matrix to form beads of about 1 .mu.m in diameter. Non-binding beads are taken away by making use of a small magnetic field in combination with a small field gradient or by rinsing with a buffer solution. The presence of the binding beads on the biochip is then detected by magnetising the particles in a relatively small, known, external magnetic field that is directed perpendicular to the plane of the substrate. [0007] Although the example given above is for detection of DNA, also other molecules such as e.g. proteins can be detected by means of the prior art BARC biochip. [0008] In the above-mentioned articles, the presence of particles is detected by making use of giant magnetoresistive (GMR) half Wheatstone bridge type sensors in the substrate, with a resistance versus applied field curve as shown in FIG. 2. The half Wheatstone bridge consists of one sensitive part above which beads are present, and one reference part above which no beads are present. The resistance versus field curve of the GMR material used is almost symmetric around zero field, as shown in FIG. 2, so that the sign of the field direction is not measured. The resistance of the GMR material decreases by about a same amount in response to a positive or negative applied field. It can be seen from FIG. 2 that there is a certain hysteresis in the GMR material, which is particularly manifest close to zero field. Consequently, accurate detection of small magnetic fields is almost impossible. [0009] The BARC biochip concept works, but the results given in FIG. 9 of D. R. Baselt, "A biosensor based on magnetoresistance technology", Biosensors & Bioelectronics 13, 731-739 (1998) show a poor signal-to-noise ratio (SNR). The main problem is that the large (1 .mu.m scale) beads used diffuse slowly through the solution, so even after a relatively long time allowed for binding between the beads and the sample DNA only a relatively small number of beads will have bound to hybridized sample molecules, leading to a weak signal. Secondly, the beads have a certain distribution of their magnetic moment (at a given field), which negatively affects the signal-to-noise ratio when only one or a few beads are present per sensor. As shown by the authors, the signal-to-noise ratio for measurement of a single bead could be enhanced by making use of smaller sensor surface areas. However, when many sensors per probe are used the electronic circuitry that is required becomes very complex. Furthermore, the slow Brownian motion of the large magnetic particles of about 1 .mu.m means that it can take a long time before the magnetic particle reaches a binding site. Thus actual measurements take a long time. [0010] Tondra et al. describe in "Model for detection of immobilized superparamagnetic nanosphere assay labels using giant magnetoresistive sensors", J. Vac. Sci. Technol. A 18(4), July/August 2000, pp. 1125-1129, that a GMR sensor can detect a single paramagnetic bead of any size, as long as a certain conditions are met, one of which is that the sensor is about the same size as the bead. This condition is easily met at a bead radius of 500 nm. Reducing the bead radius to 100 nm is possible by overcoming technical difficulties in fabrication of GMR sensors. Reducing the bead radius further to 10 nm is said to require advances in bead fabrication technology as well as in GMR sensor fabrication. A disadvantage of this solution is the required precise positioning of the probe areas with respect to the GMR sensor, on a scale well below 0.5 .mu.m. [0011] Chemla et al. describe in the article "Ultrasensitive magnetic biosensor for homogeneous immunoassay", PNAS, Dec. 19, 2000, vol. 97, no. 26, a SQUID based sensor using magnetic nanoparticles. An in-plane magnetic field is applied to de-randomise the magnetic moments of the magnetic nanoparticles attached to an immobilised zone on a substrate. The immobilised zone lies in a well and a Mylar.RTM. sheet is described as an example thereof. Then the field is switched off. The relaxation of the magnetic dipoles of the attached nanoparticles according to the Neel mechanism produces a measurable time dependence of the magnetic flux through the SQUID for a period of several seconds. This flux is detected by a SQUID probe placed close to the edge of the immobilised zone. Superparamagnetic nanoparticles in the bulk liquid are free to move according to Brownian motion and produce, in the absence of an applied field, no magnetic field. SQUID flux detectors have the disadvantage that they are expensive and that they operate only at cryogenic temperatures. [0012] It is an object of the present invention to provide a method and device for accurate detection of magnetic particles in biochips with an enhanced signal to noise ratio. [0013] It is another object of the present invention to provide a fast method for detection of magnetic particles in biochips and a corresponding device. [0014] It is still another object of the present invention to provide a method and device for detection of magnetic particles, which are simple and economical, and in particular which do not require a precise positioning of individual magnetic beads with regard to the sensors. [0015] The above objectives are accomplished, according to the present invention, by a magnetoresistive sensor device for determining the presence or an areal density of magnetic nanoparticles being directly or indirectly coupled to the target, the magnetoresistive sensor device comprising a substrate having attached thereto a binding site able to selectively bind a target, and a magnetoresistive sensor for detecting the magnetic field of magnetic nanoparticles at least when coupled to the target, wherein the magnetoresistive sensor comprises pairs of first and second magnetoresistive sensing elements or first and second groups of magnetoresistive sensing elements, each pair being associated with and located parallel with a probe element having at least one binding site, the outputs of the first and second magnetoresistive elements or first and second groups of magnetoresistive sensing elements being fed to a comparator circuit. [0016] The present invention also includes a method for determining the presence or for measuring an areal density of magnetic nanoparticles on a substrate, comprising the steps of: [0017] binding a target to selective binding sites on the substrate, the target being directly or indirectly labeled with magnetic nanoparticles, [0018] sensing the presence of the bound magnetic nanoparticles to a binding site to thereby determine the presence or density of the target labeled with magnetic nanoparticles [0019] wherein the sensing step is carried out by extracting two signals derived from the magnetic field generated by nanoparticles bound to the one binding site using magentoresistive sensors elements; and determining the difference between the two signals. [0020] The width and length dimensions of the probe areas, that are the areas on the chip at which the probe elements such as antibodies are attached, and of the magneto-resistive (MR) sensor elements, are much larger than the diameter of the magnetic nanoparticles of which the presence and concentration is to be measured. The nanoparticles may for example have a diameter between 1 and 250 nm, preferably between 3 and 100 nm, most preferred between 10 and 60 nm. For such small particles, the diffusion is fast. The width and length dimensions of sensor elements are at least a factor 10 or more, preferably a factor 100 or more, larger than the diameter of the nanoparticles, for example 1 .mu.m.times.1 .mu.m. Other dimensions for the sensor elements are also possible. If different dimensions are used, different S/N ratios are obtained. [0021] The term "micro-array" or "biochip" refers to generated arrays on a planar surface that constitute a plurality of discrete reaction or incubation compartments identifiable by their locations or x-y coordinates on the array. Such arrays are suitable for use in assays for assessing specific binding characteristics between members of specific binding pairs. The invention is very suitable in competitive assays or displacement assays. These and other features and advantages of the present invention will become apparent from the following detailed description, taken in conjunction with the accompanying drawings, which illustrate, by way of example, the principles of the invention. This description is given for the sake of example only, without limiting the scope of the invention. The reference figures quoted below refer to the attached drawings. [0022] FIG. 1 is a schematic diagram of a BARC chip according to the prior art. [0023] FIG. 2 is a graph of the response (resistance) of a multilayer GMR sensor device to an applied field according to the prior art. [0024] FIG. 3 is a perspective view of a biochip. Continue reading... Full patent description for Sensor and method for measuring the areal density of magnetic nanoparticles on a micro-array Brief Patent Description - Full Patent Description - Patent Application Claims Click on the above for other options relating to this Sensor and method for measuring the areal density of magnetic nanoparticles on a micro-array patent application. ###  How KEYWORD MONITOR works... a FREE service from FreshPatents1. Sign up (takes 30 seconds). 2. Fill in the keywords to be monitored. 3. Each week you receive an email with patent applications related to your keywords. Start now! - Receive info on patent apps like Sensor and method for measuring the areal density of magnetic nanoparticles on a micro-array or other areas of interest. ### Previous Patent Application: Diagnostic test using gated measurement of fluorescence from quantum dots Next Patent Application: Fabrication of a ferromagnetic inductor core and capacitor electrode in a single photo mask step Industry Class: Chemistry: analytical and immunological testing ### FreshPatents.com Support Thank you for viewing the Sensor and method for measuring the areal density of magnetic nanoparticles on a micro-array patent info. IP-related news and info Results in 4.30068 seconds Other interesting Feshpatents.com categories: Software: Finance , AI , Databases , Development , Document , Navigation , Error |
||
