The assignee for this patent, patent number 8641974, is
Reporters obtained the following quote from the background information supplied by the inventors: "The invention relates to a device for detecting particles of a fluid, in particular for magnetically detecting the particles.
"By immunofluorescence, marked particles, particularly also cell populations, can be analyzed and separated using a FACS system (FACS: Fluorescence Activated Cell Sorting/flow cytometry) (
"In addition to FACS separation, systems based on immunomagnetic principles are described in the literature, the emphasis here being mainly on the detection of marked cells and less on separation. Macroscopic dipole and quadrupole separators are used, as well as SQUIDS.sup.1 (Superconducting Quantum Interference Device). The disadvantages of these magnetic approaches are their impracticability (no miniaturization possible) and the high costs. Dipole and quadrupole systems, like FACS sorters, are complex and expensive instruments which, because of their size, exhibit only low marked cell recovery rates. SQUIDS must be cooled to below 100 K to be operational. In addition, the cooling necessitates a complex detector design which undermines the high sensitivity of SQUIDS.
"In modern cell separation systems, MACS (Magnetic Activated Cell Sorting) is used. Here paramagnetic nanoparticles coated with monoclonal antibodies are mixed with a cell suspension. The antibodies bind to the specific antigen on the cell surface. If the cell suspension passes through a powerful magnetic field in a column, the cell/nanoparticle complexes remain in the column, while the free cells momentarily flow through (negative selection). If the column is removed from the magnetic field, the cell/nanoparticle complexes can be recovered (positive selection). Cells attaching to microbeads are viable and the bead/antibody complex can be removed from the cell surface. MACS is completely FACS-compatible. Following magnetic separation (100.times. accumulation of up to 10.sup.9 cells in 15 min).sup.2, flow cytometry analysis with fluorescence marking can take place. The disadvantage of this process is the need for double marking (magnetic and fluorescence) in order to perform cost-effective analysis/refined separation of the cells."
In addition to obtaining background information on this patent, NewsRx editors also obtained the inventors' summary information for this patent: "One possible object is therefore to overcome the disadvantages of the related art and to create an inexpensive and accurate device and a method for detecting magnetically labeled particles, particularly cells.
"The inventors propose a device for dynamically detecting and selecting magnetically labeled particles, comprising a microfluidic channel and, disposed around same, a Wheatstone bridge circuit with at least one magnetoresistive element, the microfluidic channel being disposed in the bridge circuit such that the magnetically labeled particles flowing through the microfluidic channel measurably influence the balance of the bridge circuit.
"The inventors also propose a method for dynamically detecting and selecting magnetically labeled particles, wherein magnetically labeled particles flow through a microfluidic channel and, due to the magnetic stray field generated by them, influence at least one magnetoresistive resistor of the Wheatstone bridge disposed around the microfluidic channel so as to produce a measurable deflection of the bridge.
"Advantageously, at least two types of magnetically labeled particles are used, namely permanently magnetized particles whose dipole moment remains even after removal of an external magnetic field required for the magnetization, and temporarily magnetically labeled particles, e.g. so-called superparamagnetic particles, particularly also nanoparticles. These particles are temporarily magnetized by an external magnetic field upstream of the device and the magnetization decays with a characteristic decay time as a function of the superparamagnetic particle. The difference in the decay time enables the dynamic detection and selection described here to be carried out.
"The last mentioned magnetically marked particles are particularly preferred for the dynamic detection and selection described here.
"Ferromagnetic particles, on the other hand, are not preferred, as there is a risk of them forming clumps and, in this approach, interfering with the sensor or blocking the microfluidic channel.
"Stray field means that the field generated by the magnetically labeled particles has been induced by an external field.
"Superparamagnetic particles are produced by magnetically polarizing them in the external field. If the field is removed, with a certain decay time no dipole remains.
"The dynamic measurement is performed using a microfluidic channel. The microfluidic channel is matched to the size of the examined particles, particularly cells, in order to enable individual detection with high recovery rate. According to a preferred embodiment, the microfluidic channel is formed using a micromechanical method, e.g. by etching of a silicon wafer. The microfluidic channel is preferably matched such that each particle or each cell flows individually through the microfluidic channel, i.e. there is no room in the channel for a plurality of adjacent particles. For example, the microfluidic channel can also be produced by a forming technique such that it is made e.g. by pouring a silicone over a silicon punch. The microfluidic channel is preferably made of transparent material.
"In addition to dynamic detection and selection, another optical, electrical, magnetic and/or other detection method can also be used to analyze the fluid in the microchannel.
"The detection possibilities:
"All the cells in a microfluidic channel are detected by microscopy, absorption or fluorescence measurement and the fraction of marked cells is calculated by comparison measurement using a magnetoresistive device.
"Similarly to optical determination, the flow rate of individual cells in the microfluidic channel is counted by impedance spectroscopy, capacitive, resistive or dielectrophoretic measurement methods and the fraction of magnetically marked cells is again calculated by differential measurement with the registered number of marked cells by measurement using a magnetoresistive detector. c) Magnetic (1 type of magnetic nanoparticle): Each cell flowing through the microfluidic channel--matched to the cell dimension--is forced into the aqueous solution in the region of the magnetoresistive sensor because of the cell volume and therefore fewer unbound magnetic nanoparticles will be present. This temporary nanoparticle depletion results in a reduced measurement signal and permits the detection of unmarked cells. In the case of a marked cell, a large measurement signal is detected, as the magnetic nanoparticles accumulate on the cell surface or inside the cell. d) Magnetorelaxometry: The decay of the magnetic moment of nanoparticles is determined by the size of the particle itself and by the binding to other particles or substrates. Valuable additional information can be obtained by determining the different relaxation times by time-resolved measurements with the magnetoresistive sensor. For example, bound and unbound cells can be differentiated. It is conceivable to provide various cell types or cells with different antigen bonds and bind respective nanoparticles with different size sorting. Different sized cells as ligands to the same kind of magnetic nanoparticles can also be differentiated in this way.
"The proposed solution is based on the use of magnetoresistive components which are incorporated in the microfluidic channels. Unlike in all immunomagnetic approaches hitherto, cell detection takes place dynamically during the flow of cells through a microfluidic channel. The setup is schematically illustrated in the accompanying drawings. A Wheatstone bridge arrangement of generally 4 magnetoresistive resistors is disposed such that two resistors are mounted below a microfluidic channel and two resistors remain outside the microfluidic region in order to achieve maximum unbalancing of the bridge by magnetically marked cells. However, the arrangement of the 4 elements is variable and, depending on the arrangement, the signal sensitivity or local resolution of the device and of the method can be optimized.
"Moreover, there do not necessarily have to be 4 magnetoresistive elements in the bridge circuit. It is equally possible for 2 magnetoresistive elements to be combined with two conventional resistors in the bridge. Or any other combination, as long as at least one magnetoresistive element is present in the bridge circuit.
"The magnetoresistive elements can be both conventional magnetoresistive elements, as described e.g. in DE 102 02 287, or organic magnetoresistive elements as disclosed, for example, in DE 10 2006 019 482.
"Analysis and separation of particles, e.g. heterogeneous cell populations, on the basis of cellular features is accessible by magnetoresistive measurements. For the separation of the heterogeneous cell populations, immunomagnetic markers (nanoparticles bound to a receptor or ligand) are used. The marked cells are magnetized in an external magnetic field, thereby generating an additional magnetic stray field which can be detected by a magnetoresistive component. Through the use of magnetorelaxometry (decay of the stray field over time), unbound and different bound magnetic nanoparticles can be differentiated from one another. No other fluorescence markings or similar are necessary for the separation or analysis of the cell population.
"The marked cells are detected by a magnetoresistive component, whereas the unmarked cells can be optically, electrically or magnetoresistively detected.
"The proposed device and method have the potential to replace fluorescence measurement in conventional FACS analysis. For this type of fluorescence-based cell detection mentioned in the introduction, the cells are introduced into a small capillary tube. Fluorescence detection takes places discretely on individual cells. The individual cells are atomized into small droplets and electrically charged as a function of fluorescence detection. The charged droplets are deflected in a collector and collected in a vessel.
"The GMR sensor is here inserted upstream of the atomizer. Measurements into the MHz range can be performed with this setup. The advantage of this design is its massive potential for miniaturization and parallelization. In addition to separation using an atomizer, analysis of the cell population can also be performed.
"In hitherto known applications of GMR sensing in biology and medicine (DNA sensor), immobilized, magnetically marked material is detected, a statistical selection method being used. For the dynamic measurement of individual analytes, high-frequency interrogation of the sensor status is advantageous. This is implemented e.g. by field scanning of the sensor in the relevant region of the characteristic at frequencies typically significantly higher than 1 kHz. A deviation in the sensor amplitude or a step-change in the sensor sensitivity (differentiated signal) are e.g. an indication of event detection.
"This selection method additionally allows electronic lock-in filtering to be used for conditioning of the wanted signal, thereby enabling event detection sensitivity to be significantly increased.
"The magnetoresistive sensor is advantageously implemented by disposing at least one magnetoresistive element in the form of a Wheatstone bridge. For dynamic detection of magnetic particles, the elements of the bridge are spaced at least the particle size apart. The bridge then has an effective extent of four particle diameters. This means that detecting closely consecutive particles becomes a problem. Assuming that the fluidic channel can be precisely microfabricated and positioned on a carrier chip with a high degree of accuracy, the geometry of the bridge circuit can be modified such that two diagonal elements of the Wheatstone geometry lie inside and the two other diagonal elements outside the microfluidic cell, thereby doubling the local resolution for particle detection.
"In another exemplary embodiment, three bridge elements are placed outside the detection region. Although this halves the signal sensitivity of the sensor, the local resolution is quadrupled."
For more information, see this patent: Bar,
Keywords for this news article include: Antibodies, Immunology, Blood Proteins, Nanotechnology, Immunoglobulins, Emerging Technologies, Magnetic Nanoparticles,
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