EP2404155A1 - Device and method for concentrating and detecting magnetically marked cells in laminarly flowing media - Google Patents

Device and method for concentrating and detecting magnetically marked cells in laminarly flowing media

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Publication number
EP2404155A1
EP2404155A1 EP10707007A EP10707007A EP2404155A1 EP 2404155 A1 EP2404155 A1 EP 2404155A1 EP 10707007 A EP10707007 A EP 10707007A EP 10707007 A EP10707007 A EP 10707007A EP 2404155 A1 EP2404155 A1 EP 2404155A1
Authority
EP
European Patent Office
Prior art keywords
cells
magnetic field
magnetoresistor
detection
channel
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP10707007A
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German (de)
French (fr)
Inventor
Oliver Hayden
Manfred Rührig
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Siemens AG
Original Assignee
Siemens AG
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Filing date
Publication date
Application filed by Siemens AG filed Critical Siemens AG
Publication of EP2404155A1 publication Critical patent/EP2404155A1/en
Withdrawn legal-status Critical Current

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Classifications

    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B03—SEPARATION OF SOLID MATERIALS USING LIQUIDS OR USING PNEUMATIC TABLES OR JIGS; MAGNETIC OR ELECTROSTATIC SEPARATION OF SOLID MATERIALS FROM SOLID MATERIALS OR FLUIDS; SEPARATION BY HIGH-VOLTAGE ELECTRIC FIELDS
    • B03C—MAGNETIC OR ELECTROSTATIC SEPARATION OF SOLID MATERIALS FROM SOLID MATERIALS OR FLUIDS; SEPARATION BY HIGH-VOLTAGE ELECTRIC FIELDS
    • B03C1/00—Magnetic separation
    • B03C1/02—Magnetic separation acting directly on the substance being separated
    • B03C1/28—Magnetic plugs and dipsticks
    • B03C1/288—Magnetic plugs and dipsticks disposed at the outer circumference of a recipient
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
    • B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip
    • B01L3/502761—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip specially adapted for handling suspended solids or molecules independently from the bulk fluid flow, e.g. for trapping or sorting beads, for physically stretching molecules
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B03—SEPARATION OF SOLID MATERIALS USING LIQUIDS OR USING PNEUMATIC TABLES OR JIGS; MAGNETIC OR ELECTROSTATIC SEPARATION OF SOLID MATERIALS FROM SOLID MATERIALS OR FLUIDS; SEPARATION BY HIGH-VOLTAGE ELECTRIC FIELDS
    • B03C—MAGNETIC OR ELECTROSTATIC SEPARATION OF SOLID MATERIALS FROM SOLID MATERIALS OR FLUIDS; SEPARATION BY HIGH-VOLTAGE ELECTRIC FIELDS
    • B03C1/00—Magnetic separation
    • B03C1/02—Magnetic separation acting directly on the substance being separated
    • B03C1/025—High gradient magnetic separators
    • B03C1/031—Component parts; Auxiliary operations
    • B03C1/033—Component parts; Auxiliary operations characterised by the magnetic circuit
    • B03C1/0335—Component parts; Auxiliary operations characterised by the magnetic circuit using coils
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/40—Concentrating samples
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0861—Configuration of multiple channels and/or chambers in a single devices
    • B01L2300/0877—Flow chambers
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/04—Moving fluids with specific forces or mechanical means
    • B01L2400/0403—Moving fluids with specific forces or mechanical means specific forces
    • B01L2400/043—Moving fluids with specific forces or mechanical means specific forces magnetic forces
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B03—SEPARATION OF SOLID MATERIALS USING LIQUIDS OR USING PNEUMATIC TABLES OR JIGS; MAGNETIC OR ELECTROSTATIC SEPARATION OF SOLID MATERIALS FROM SOLID MATERIALS OR FLUIDS; SEPARATION BY HIGH-VOLTAGE ELECTRIC FIELDS
    • B03C—MAGNETIC OR ELECTROSTATIC SEPARATION OF SOLID MATERIALS FROM SOLID MATERIALS OR FLUIDS; SEPARATION BY HIGH-VOLTAGE ELECTRIC FIELDS
    • B03C2201/00—Details of magnetic or electrostatic separation
    • B03C2201/26—Details of magnetic or electrostatic separation for use in medical applications

Definitions

  • the invention relates to a device and a method for enrichment and detection of cells in flowing media, in particular of labeled cells in complex media such as blood. 10
  • analytes can also be sorted to a limited extent according to size and magnetic moment, see N. Pamme and A. Manz, Anal. Chem., 2004, 76, 7250.
  • magnetoresistive sensor manufacturers are only offering assays for DNA and protein analysis for in vitro diagnostic use. For example, refer to the Internet addresses of magnabiosciences.com, diagnsticbioseners.com, seahawkbio.com and san.rr.com/magnesensors.
  • a magnetoresistive sensor e.g., GMR sensor
  • GMR sensor magnetoresistive sensor
  • the object of the present invention is therefore to overcome the disadvantages of the prior art and to provide an apparatus and a method for single-cell detection in the flowing medium.
  • the subject matter of the invention is an apparatus for enrichment and detection of cells, wherein at least one magnetoresistor is arranged in an outer, surrounding magnetic field below a channel in which a laminar flow of a medium flows with magnetically marked cells.
  • the invention relates to a method for enrichment and detection of magnetically labeled cells in a laminar flowing medium, wherein cells on a
  • Magnetoresistance be enriched by an external magnetic field.
  • the invention discloses the technique of obtaining an accumulation of labeled cells directly on the magnetoresistors by means of an external magnetic field so that a nearly 100% detection of the labeled cells can be achieved.
  • the flow cytometry shown here makes it almost possible to enumerate individual labeled cells with a near 100% recovery rate when the GMR component overflows dynamically in the flowing medium.
  • treacherous diseases such as cancer, it is sometimes necessary that in about 10ml of whole blood 1 to 100 cells are quantifiable.
  • Single labeled cells in a complex matrix such as blood or partially purified (typically 1: 1000 to 1: 1 000 000) while in the flowing medium are directed to the substrate surface (as close as possible to the GMR sensor)
  • the magnetic field is applied so that an amplification of the gradient of the magnetic field directly below the GMR sensors takes place so that the entry point of the magnetic field lines in the sample space is as close as possible to the GMR sensors.
  • the magnet is arranged directly below the GMR sensors.
  • the embodiment in which the magnet for the outer, the magnetoresistors surrounding magnetic field is beveled on one or both sides, so that results in a flux concentration and an increase of the magnetic field gradient.
  • Cell detection with magnetoresistors is most easily carried out with sophisticated sensors such as the AMR, GMR and / or TMR sensors, the two latter being advantageously designed as so-called spin valves.
  • the laminar flow causes the cells to be transported without turbulence in the liquid stream.
  • cells that come in contact with the surface are caused to rotate due to shear forces and the airfoil.
  • the effect is exploited to on the one hand as possible lead all labeled cells to the magnetoresistors, and on the other hand introduce the statistically distributed immunomagnetic marker on the cell surface by "rolling" close to the GMR sensors.
  • the accumulation of cells in a magnetic field gradient which is currently used only for cell separation, is suitable for specifically enriching the labeled cells from the laminar flow to the substrate surface with the magnetoresistors depending on flow rate and number of magnetic labels per cell.
  • the magnetic force and thus the shear force or holding force can be varied to the enriched cells without the transport of unlabelled cells along the microfluidic channel to prevent.
  • this measurement target is achieved by the following components of a measurement system interacting:
  • GMR sensor has a dimension equivalent to the diameter of a single cell (typically 5-40 microns), to achieve high signal-to-noise ratio and to detect signal from only one cell
  • An external magnetic gradient field is preferably used to guide stochastically distributed and labeled cells in a microfluidic channel to the substrate surface (typically the cell to GMR sensor distance will be 0-1 ⁇ m); then the signal-to-noise ratio can be increased
  • the flowing medium is advantageously laminar, since turbulence can lead to a reduction in the recovery rate of labeled cells.
  • Typical channels have a cross section of 100-1000 ⁇ m width and 100-1000 ⁇ m height. This means that a GMR sensor with cell dimensions is much smaller than the channel size.
  • the individual labeled cells are controlled in the immediate vicinity of the substrate in the flowing medium out.
  • a stochastic distribution of labeled cells on the substrate surface leads to counting losses (for example at 10 ⁇ m GMR in a 100 ⁇ m channel ⁇ 90% loss). Cells are therefore guided along, for example, ferromagnetic strips directly onto a sensor.
  • This measuring arrangement also has the advantage that in the ideal case only a single GMR sensor is necessary to count all marked cells.
  • FIG. 1 shows the two cross sections through an embodiment of a microfluidic channel according to the invention, on the left a cross section along the flow direction and on the right a cross section perpendicular to the flow direction.
  • Figure 1 shows schematically the process of cell enrichment on the substrate surface 8 with the GMR sensors.
  • FIG 1 shows a longitudinal cross section of a microfluidic channel 4 in which a laminar flow, as indicated by the arrow 5, is shown in the left part of the FIGURE. flows.
  • a laminar flow as indicated by the arrow 5
  • unlabeled cells 2 which move evenly distributed in the laminar flow.
  • a magnet 7 is arranged; the image immediately shows the accumulation of the marked cells on the bottom / substrate 8 of the channel within the magnetic field gradient 7.
  • the GMR sensors like all magnetoresistors, can also be connected to the Side walls of the channel wall and / or be arranged at the top of the channel.
  • the enrichment of cells with superparamagnetic markers 1 from a complex medium in a magnetic field 9 is shown.
  • the laminar flow 5 prevents swirling of the cells 1 and 2.
  • the cells 1,2 can roll along the substrate surface 8 and thus come into closest contact with the GMR sensors 3.
  • the strength of the magnetic field but the transport does not interfere with the labeled cells in the microfluidic channel, which can be adjusted, for example, by suitable pulsed operation and by the symmetry of the gradient field.
  • the microfluidic channel 4 can be seen in cross-section through the flow direction.
  • the field lines 9 of the magnetic field which have their origin in the GMR sensors 3 and therefore cause a gradient amplification of the magnetic field.
  • the magnet 7 has at least one bevel 6 towards the GMR sensors, but preferably 2 bevels 6 as shown.
  • Figure 2 shows the same image as Figure 1 in longitudinal cross-section and illustrates the cell rolling within the laminar flow 5.
  • the three phases of the CeIl- Rollings can be seen, first (A) the enrichment of the labeled cells 1 on the substrate surface of the bottom. 8 of the microfluidic channel 4 in the magnetic field 9, then (B) cell rolling over the sensor surface, where (C) cell detection takes place.
  • the gradient magnetic field (-100 mT with dB / dx of a few 10-100 T / m, depending on the loading of the cells with superparamagnetic particles) pulsed.
  • the detection of the labeled cells takes place in a weak measuring magnetic field of ⁇ ImT.
  • FIG. 3 shows in time sequence the strength of the magnetic field for cell enrichment, cell detection and GMR measurement.
  • the time is plotted on the X-axis, so that it can be seen that always two magnetic field strengths are applied in temporal change.
  • a method of continuous cell enrichment and cell detection may be performed by a sequence of pulsed magnetic fields.
  • FIG. 3 shows the cyclical sequence of (1) enrichment, (2) + (3) measurement for a continuous measurement, which is illustrated graphically.
  • the measurement and accumulation of the cells can thus be tracked or controlled independently of each other in the kHz range.
  • Figure 3 how at the very top with a "strong magnetic field and long pulse times" cell enrichment takes place within the microfluidic channel, below which is a graph showing that a weaker magnetic field with less pulse time is used for cell detection Graphic as with low magnetic field and short pulse time the GMR measurement is completed.
  • FIG. 4 again shows a microfluidic channel, again in cross section perpendicular to the flow direction as in FIG. 1 on the right side.
  • the measuring magnetic field can be applied vertically or in the same plane as the GMR sensors (FIG. 4).
  • the magnet (magnetic yoke) of the gradient magnetic field can be used to set a gradient in the measuring magnetic field in order to achieve a local detuning of the bridge members of the GMR measuring bridge. This detuning represents the measurement signal for the concentration of the magnetic particles in the sensor region.
  • the measuring magnetic field can additionally be time-modulated in order, for example, to obtain a time delay. using lock-in technology and suppressing the low-frequency noise (1 / f noise) to improve the signal-to-noise ratio.
  • pulsed magnetic fields are enriched and detected as shown in FIG.
  • FIG. 4 the schematic arrangement of the magnets or coils 7, 10 and 11 for enrichment and detection around the microfluidic channel 4 is shown.
  • the magnet 7 for the strong magnetic field is applied for enrichment below the GMR sensors and the coils 10 and 11 for the weak magnetic field are applied perpendicular to the GMR sensor for detection. Both fields can be controlled separately with 2 magnets, wherein preferably the weak magnetic field is applied in the sensor plane.
  • the accumulation of the cells or the applied shear force on the cells can be controlled by the magnetic field strength and the flow rate.
  • Marked cells are near the surface and can be sensitively detected with magnetoresistive components.
  • the presented method allows a large area application for multiplexing (e.g., array of GMR sensors).
  • Cell rolling can be adapted to the application by means of surface-functionalized microfluidic channels, for example functionalized with receptors (selectins), biological components (proteins, polysaccharides), or SAMs (self-assembled mono- layer) or by silanization.
  • receptors selectiveins
  • biological components proteins, polysaccharides
  • SAMs self-assembled mono- layer
  • Circulating tumor cells CTC
  • tumor stem cells inflammatory cells
  • stem cells inflammatory cells
  • bacteria or yeasts may precede the actual detection in the flowing medium.
  • the magnetic detection can be done with optical methods
  • Areas of application in the human area include:
  • Oncology regenerative medicine, infectiology, clinical diagnostics, clinical chemistry, imaging.

Abstract

The invention relates to a device and to a method for concentrating and detecting cells in flowing media, in particular magnetically marked cells in complex media such as blood. For this purpose, at least one magnet is used, said magnet being coupled to at least one magnetoresistance.

Description

Beschreibung description
Vorrichtung und Verfahren zur Anreicherung und Erfassung von Zellen in strömenden Medien 5Device and method for enrichment and detection of cells in flowing media 5
Die Erfindung betrifft eine Vorrichtung und ein Verfahren zur Anreicherung und Erfassung von Zellen in strömenden Medien, insbesondere von markierten Zellen in komplexen Medien wie beispielsweise Blut. 10The invention relates to a device and a method for enrichment and detection of cells in flowing media, in particular of labeled cells in complex media such as blood. 10
Bisher gibt es keine nicht-optische Lösung um verlässliche Einzelzelldetektion mit magnetischen Durchflusszytometer in laminarer Strömung durchzuführen.So far, there is no non-optical solution to perform reliable single cell detection with magnetic flow cytometer in laminar flow.
15 Die gegenwärtig bekannten technischen Lösungen zur Einzelzelldetektion sind vorwiegend optische Methoden, um Zellen mit fluoreszierenden Markern oder durch Streulicht aus einer Suspension in Durchflusskanälen zu detektieren. Magnetische Methoden beschränken sich vorwiegend auf Anreicherungen vonThe presently known technical solutions for single-cell detection are predominantly optical methods for detecting cells with fluorescent markers or scattered light from a suspension in flow channels. Magnetic methods are mainly limited to enrichments of
20 magnetisch markierten Zellen sowie Biosensoren mit magnetore- sistiven Transducer.20 magnetically labeled cells and biosensors with magnetoresistive transducers.
Folgende magnetische Methoden sind bekannt:The following magnetic methods are known:
251) Anlegen eines externen Magnetfeldes rechtwinklig zur Durchflussrichtung. In einem Gradientenfeld können zudem im eingeschränkten Masse Analyte nach Größe und magnetischem Moment sortiert werden, siehe dazu N. Pamme and A. Manz, Anal. Chem., 2004, 76, 7250.251) Applying an external magnetic field perpendicular to the flow direction. In a gradient field, analytes can also be sorted to a limited extent according to size and magnetic moment, see N. Pamme and A. Manz, Anal. Chem., 2004, 76, 7250.
3030
2) Einbau eines ferromagnetischen Leiters im Boden der Trennkammer. Durch den lokalen Feldgradienten werden magnetisier- bare Zellen am Boden entlang des ferromagnetischen Leiters angereichert bei Durchflussraten von <1 mm/s von unmarkierten2) Installation of a ferromagnetic conductor in the bottom of the separation chamber. Due to the local field gradient, magnetizable cells are enriched at the bottom along the ferromagnetic conductor at flow rates of <1 mm / s unlabeled
35 Zellen getrennt D. W. Inglis, R. Riehn, R. H. Austin and J. C. Sturm, Appl. Phys . Lett., 2004, 85, 5093. 3) Ein Stromleiter wird am Boden der Trennkammer eingebracht. Durch den Stromfluss wird ein Magnetfeld induziert, dass wiederum - wie unter Punkt 2 ausgeführt - verwendet werden kann, um Zellen anzureichern (Durchflussrate: 6 nl/min in Mikrofluidikkanälen) Pekas, N., Granger, M., Tondra,35 cells separated DW Inglis, R. Riehn, RH Austin and JC Sturm, Appl. Phys. Lett., 2004, 85, 5093. 3) A conductor is placed at the bottom of the separation chamber. The current flow induces a magnetic field which, in turn, can be used to enrich cells (flow rate: 6 nl / min in microfluidic channels) as described under point 2) Pekas, N., Granger, M., Tondra,
M., Popple, A. and Porter, M. D, Journal of Magnetism and Magnetic Materials, 293, pp . 584-588, (2005) . c) M. Tondra, M. Granger, R. Fuerst, M. Porter, C. Nordman, J. Taylor, and S. Akou, IEEE Transactions on Magnetics 37, (2001), pp . 2621- 2623.M., Popple, A. and Porter, M.D., Journal of Magnetism and Magnetic Materials, 293, pp. 584-588, (2005). c. M. Tondra, M. Granger, R. Fuerst, M. Porter, C. Nordman, J. Taylor, and S. Akou, IEEE Transactions on Magnetics 37, (2001), pp. 2621-2623.
Die Detektion markierter Zellen mit eingebetteten GMR Sensoren kann bisher nur statisch analog zu einem Assay und nicht dynamisch, also beispielsweise in laminarer Strömung, durch- geführt werden. siehe dazu: J. Schotter, P. B. Kamp, A. Becker, A. Puhler, G. Reiss and H. Brückl, Biosens. Bioe- lectron., 2004, 19, 1149.The detection of labeled cells with embedded GMR sensors can hitherto only be carried out statically analogously to an assay and not dynamically, ie, for example, in a laminar flow. see: J. Schotter, P. B. Kamp, A. Becker, A. Puhler, G. Reiss and H. Brückl, Biosens. Bioelectronics., 2004, 19, 1149.
Kommerzielle Hersteller von Sensoren mit magnetoresistiven Elementen bieten nur Assays für DNA und Proteinanalytik für die In-vitro-Diagnostik an. Dazu sei beispielsweise auf die Internet-Adressen von magnabiosciences.com, diagnsticbiosen- sors.com, seahawkbio.com und san.rr.com/magnesensors verwiesen .Commercial magnetoresistive sensor manufacturers are only offering assays for DNA and protein analysis for in vitro diagnostic use. For example, refer to the Internet addresses of magnabiosciences.com, diagnsticbioseners.com, seahawkbio.com and san.rr.com/magnesensors.
In bekannten magnetischen Durchflusszytometer werden Zellen, die mit magnetischen, beispielsweise superparamagnetischen Labels, markiert sind, in einer Durchflusskammer oberflächennah über einen magnetoresistiven Sensor (z.B. GMR Sensor) transportiert wie z.B. N. Pekas, M. D. Porter, M. Tondra, A. Popple and A. Jander, Appl . Phys . Lett., 2004, 85, 4783 beschrieben .In known magnetic flow cytometers, cells labeled with magnetic, such as superparamagnetic, labels are transported in a flow-through chamber near the surface via a magnetoresistive sensor (e.g., GMR sensor), such as e.g. N. Pekas, M.D. Porter, M. Tondra, A. Popple and A. Jander, Appl. Phys. Lett., 2004, 85, 4783.
Problematisch dabei ist, dass die erforderliche Nähe der mar- kierten Zelle zum Sensor nicht erreicht wird, da das magnetische Streufeld durch die magnetischen Marker mit der dritten Potenz zum Abstand vom Sensor abfällt. Es können daher mit den bislang bekannten Verfahren in der Regel bei weitem nicht alle markierten Zellen erfasst werden.The problem with this is that the required proximity of the marked cell to the sensor is not reached, since the magnetic stray field is dropped by the third-power magnetic markers at a distance from the sensor. It can therefore with As a rule, not all labeled cells are detected by the methods known hitherto.
Aufgabe der vorliegenden Erfindung ist es daher, die Nachtei- Ie des Standes der Technik zu überwinden und eine Vorrichtung und ein Verfahren zur Einzelzelldetektion im strömenden Medium anzugeben.The object of the present invention is therefore to overcome the disadvantages of the prior art and to provide an apparatus and a method for single-cell detection in the flowing medium.
Diese Aufgabe wird durch den Gegenstand der Erfindung, wie er in den Ansprüchen, der Beschreibung und den Figuren offenbart ist, gelöst.This object is achieved by the subject matter of the invention as disclosed in the claims, the description and the figures.
Demnach ist Gegenstand der Erfindung eine Vorrichtung zur Anreicherung und Detektion von Zellen, wobei unterhalb eines Kanals, in dem eine laminare Strömung eines Mediums mit magnetisch markierten Zellen fließt, zumindest ein Magnetowiderstand in einem äußeren, ihn umgebenden Magnetfeld angeordnet ist. Außerdem ist Gegenstand der Erfindung ein Verfahren zur Anreicherung und Detektion von magnetisch markierten Zellen in einem laminar strömenden Medium, wobei Zellen an einemAccordingly, the subject matter of the invention is an apparatus for enrichment and detection of cells, wherein at least one magnetoresistor is arranged in an outer, surrounding magnetic field below a channel in which a laminar flow of a medium flows with magnetically marked cells. In addition, the invention relates to a method for enrichment and detection of magnetically labeled cells in a laminar flowing medium, wherein cells on a
Magnetowiderstand durch ein äußeres Magnetfeld angereichert werden .Magnetoresistance be enriched by an external magnetic field.
Die Erfindung offenbart also erstmalig die Technik, wie durch ein äußeres Magnetfeld eine Anreicherung von markierten Zellen direkt an den Magnetowiderständen erreicht wird, so dass eine nahezu 100% Erfassung der markierten Zellen erreichbar ist .Thus, for the first time, the invention discloses the technique of obtaining an accumulation of labeled cells directly on the magnetoresistors by means of an external magnetic field so that a nearly 100% detection of the labeled cells can be achieved.
Dabei handelt es sich eben um Zellen, wie sie in Lebewesen, beispielsweise Tiere/Menschen vorkommen.These are just cells, as they occur in living things, such as animals / humans.
Durch die hier gezeigte Durchflsszytometrie ist es nahezu möglich, einzelne markierte Zellen mit nahe 100% Wiederfin- dungsrate beim Überfließen des GMR-Bauteils dynamisch im fließenden Medium abgezählt werden können. Insbesondere bei tückischen Krankheiten wie Krebs ist es zum Teil nötig, dass in ca. 10ml Vollblut 1 bis 100 Zellen quantifizierbar sind.The flow cytometry shown here makes it almost possible to enumerate individual labeled cells with a near 100% recovery rate when the GMR component overflows dynamically in the flowing medium. In particular, in treacherous diseases such as cancer, it is sometimes necessary that in about 10ml of whole blood 1 to 100 cells are quantifiable.
Nach einer vorteilhaften Ausführungsform werdenAccording to an advantageous embodiment
a. Einzelne markierte Zellen in einer komplexen Matrix wie Blut oder teilaufgereinigt (typischerweise 1:1000 bis 1:1 000 000), während Sie sich im strömenden Medium befinden, an die Substratoberfläche geführt (so nah wie möglich am GMR sensor)a. Single labeled cells in a complex matrix such as blood or partially purified (typically 1: 1000 to 1: 1 000 000) while in the flowing medium are directed to the substrate surface (as close as possible to the GMR sensor)
b. An der Oberfläche werden sie hinsichtlich des GMR Sensors im laminaren Fluss ausgerichtet (Zellen dürfen nicht stochastisch verteilt über das Substrat mit dem Sensor/den Sensoren fließen)b. On the surface, they are aligned with respect to the GMR sensor in laminar flow (cells must not flow stochastically across the substrate with the sensor (s))
c. Detektion von Zellen erfolgt einzeln (werden „abgezählt"; daher magnetische Durchflusszytometrie) ; dazu ist ein ausreichend hohes Signal-Rausch-Verhältnis vorteilhaft.c. Detection of cells is done individually (counted out, therefore magnetic flow cytometry) and a high signal-to-noise ratio is advantageous.
Nach einer vorteilhaften Ausführungsform des Verfahrens wird das Magnetfeld so angelegt, dass eine Verstärkung des Gradienten des Magnetfelds direkt unterhalb der GMR-Sensoren so stattfindet, dass der Eintrittspunkt der Magnet-Feldlinien in den Probenraum möglichst nahe bei den GMR-Sensoren liegt.According to an advantageous embodiment of the method, the magnetic field is applied so that an amplification of the gradient of the magnetic field directly below the GMR sensors takes place so that the entry point of the magnetic field lines in the sample space is as close as possible to the GMR sensors.
Dazu wird nach einer vorteilhaften Ausführungsform der Vorrichtung der Magnet direkt unterhalb der GMR-Sensoren ange- ordnet.For this purpose, according to an advantageous embodiment of the device, the magnet is arranged directly below the GMR sensors.
Insbesondere vorteilhaft ist die Ausführungsform, bei der der Magnet für das äußere, die Magnetowiderstände umgebende Magnetfeld an einer oder beiden Seiten angeschrägt wird, so dass daraus eine Flusskonzentration und ein Erhöhung des Magnetfeldgradienten resultiert. Die Zelldetektion mit Magnetowiderständen erfolgt am einfachsten mit den technisch ausgereiften Sensoren wie den AMR, GMR, und/oder TMR - Sensoren, wobei die beiden letzteren vorteilhaft als sogenannte Spin Valves ausgeführt sind.Particularly advantageous is the embodiment in which the magnet for the outer, the magnetoresistors surrounding magnetic field is beveled on one or both sides, so that results in a flux concentration and an increase of the magnetic field gradient. Cell detection with magnetoresistors is most easily carried out with sophisticated sensors such as the AMR, GMR and / or TMR sensors, the two latter being advantageously designed as so-called spin valves.
Die laminare Strömung bewirkt, dass die Zellen ohne Verwirbe- lung im Flüssigkeitsstrom transportiert werden. Jedoch werden Zellen, die in Kontakt mit der Oberfläche kommen, aufgrund auftretender Scherkräfte und dem Strömungsprofil in Rotation gebracht. Gemäß der Erfindung wird der Effekt ausgenutzt, um einerseits möglichst alle markierten Zellen an die Magnetowiderstände hinzu führen, und andererseits die statistisch verteilen immunomagnetischen Marker auf der Zelloberfläche durch „Abrollen" nahe an die GMR Sensoren heranzuführen.The laminar flow causes the cells to be transported without turbulence in the liquid stream. However, cells that come in contact with the surface are caused to rotate due to shear forces and the airfoil. According to the invention, the effect is exploited to on the one hand as possible lead all labeled cells to the magnetoresistors, and on the other hand introduce the statistically distributed immunomagnetic marker on the cell surface by "rolling" close to the GMR sensors.
Die Anreicherung von Zellen in einem Magnetfeldgradienten, die gegenwärtig nur für die Zellseparation eingesetzt wird, eignet sich, um gezielt in Abhängigkeit von Durchflussrate und Anzahl der magnetischen Labels pro Zelle die markierten Zellen aus der laminaren Strömung an die Substratoberfläche mit den Magnetowiderständen anzureichern. Darüber hinaus kann durch die die Durchflussgeschwindigkeit und die Stärke des Gradienten die Magnetkraft und damit die Scherkraft bzw. Haltekraft auf die angereicherten Zellen variiert werden ohne den Transport von unmarkierten Zellen entlang des mikroflui- dischen Kanals zu unterbinden.The accumulation of cells in a magnetic field gradient, which is currently used only for cell separation, is suitable for specifically enriching the labeled cells from the laminar flow to the substrate surface with the magnetoresistors depending on flow rate and number of magnetic labels per cell. In addition, by the flow rate and the strength of the gradient, the magnetic force and thus the shear force or holding force can be varied to the enriched cells without the transport of unlabelled cells along the microfluidic channel to prevent.
Bevorzugt wird dieses Messziel dadurch erreicht, dass folgende Komponenten eines Messsystems zusammenspielen:Preferably, this measurement target is achieved by the following components of a measurement system interacting:
d. GMR Sensor hat eine Dimension, die dem Durchmesser einer Einzelzelle entspricht (typischerweise 5-40 μm) , um hohes Signal-Rausch-Verhältnis zu erzielen und Signal nur von einer Zelle zu detektierend. GMR sensor has a dimension equivalent to the diameter of a single cell (typically 5-40 microns), to achieve high signal-to-noise ratio and to detect signal from only one cell
e. Um einzelne Zellen in einem großen Probenvolumen messen zu können, werden diese in einem strömenden Medium geführt f. Ein externes magnetisches Gradientenfeld wird bevorzugt eingesetzt, um stochastisch verteilte und markierte Zellen in einem Mikrofluidikkanal an die Substratoberfläche zu führen (typischerweise wird der Abstand der Zelle zum GMR Sensor 0-1 μm betragen) ; dann kann das Signal-Rausch Verhältnis erhöht werdene. In order to measure individual cells in a large sample volume, they are guided in a flowing medium f. An external magnetic gradient field is preferably used to guide stochastically distributed and labeled cells in a microfluidic channel to the substrate surface (typically the cell to GMR sensor distance will be 0-1 μm); then the signal-to-noise ratio can be increased
g. Das strömende Medium ist vorteilhafterweise laminar, da eine Verwirbelung zur Reduktion der Wiederfindungsra- te von markierten Zellen führen kann. Typische Kanäle haben einen Querschnitt von 100- 1000 μm Breite und 100-1000 μm Höhe. Das bedeutet, dass ein GMR Sensor mit Zelldimensionen wesentlich kleiner ist als die Kanalgröße.G. The flowing medium is advantageously laminar, since turbulence can lead to a reduction in the recovery rate of labeled cells. Typical channels have a cross section of 100-1000 μm width and 100-1000 μm height. This means that a GMR sensor with cell dimensions is much smaller than the channel size.
Die einzelnen markierten Zellen werden kontrolliert in unmittelbarer Substratnähe im strömenden Medium geführt. Eine sto- chastische Verteilung von markierten Zellen auf der Substratoberfläche führt zu Zählverlusten (beispielsweise bei lOμm GMR in einem 100 μm Kanal ~90%Verlust) . Zellen werden daher entlang von beispielsweise ferromagnetischen Streifen direkt auf einen Sensor hingeführt. Diese Messanordnung hat zudem den Vorteil, dass im Idealfall nur ein einzelner GMR Sensor notwendig ist, um alle markierten Zellen abzuzählen.The individual labeled cells are controlled in the immediate vicinity of the substrate in the flowing medium out. A stochastic distribution of labeled cells on the substrate surface leads to counting losses (for example at 10 μm GMR in a 100 μm channel ~ 90% loss). Cells are therefore guided along, for example, ferromagnetic strips directly onto a sensor. This measuring arrangement also has the advantage that in the ideal case only a single GMR sensor is necessary to count all marked cells.
Im Folgenden wird die Erfindung noch anhand einiger Figuren näher erläutert:In the following, the invention will be explained in more detail with reference to a few figures:
Figur 1 zeigt die zwei Querschnitte durch eine Ausführungsform eines Mikrofluidkanals gemäß der Erfindung, links einen Querschnitt entlang der Strömungsrichtung und rechts einen Querschnitt senkrecht zur Strömungsrichtung.FIG. 1 shows the two cross sections through an embodiment of a microfluidic channel according to the invention, on the left a cross section along the flow direction and on the right a cross section perpendicular to the flow direction.
Figur 1 zeigt schematisch den Prozess der Zellanreicherung auf der Substratoberfläche 8 mit den GMR-Sensoren .Figure 1 shows schematically the process of cell enrichment on the substrate surface 8 with the GMR sensors.
Zu erkennen ist ein im linken Teil der Figur ein längsseitiger Querschnitt eines Mikrofluidikkanals 4 in dem eine laminare Strömung, wie sie über den Pfeil 5 angezeigt wird, strömt. Dabei befinden sich in der Nähe zum Pfeil 5 markierte Zellen 1 und unmarkierte Zellen 2, die sich gleichmäßig verteilt in der laminaren Strömung bewegen. Etwas rechts davon und unterhalb des Mikrofluidikkanals 4 ist ein Magnet 7 ange- ordnet, man erkennt im Bild sofort die Anreicherung der markierten Zellen am Boden/Substrat 8 des Kanals innerhalb des Magnetfeldgradienten 7. Die GMR-Sensoren wie alle Magnetowiderstände können dabei auch an den Seitenwänden der Kanalwand und/oder oben am Kanal angeordnet sein. Wiederum etwas weiter rechts, also in Strömungsrichtung, befinden sich am Boden/Substrat 8 des Kanals mehrere GMR Sensoren 3. Durch das „Cell-Rolling" am Kanalboden und die Anreicherung der markierten Zellen durch das äußere Magnetfeld ist es möglich, dass möglichst viele der markierten Zellen auch tatsächlich von den GMR-Sensoren erfasst werden.1 shows a longitudinal cross section of a microfluidic channel 4 in which a laminar flow, as indicated by the arrow 5, is shown in the left part of the FIGURE. flows. In this case, in the vicinity of the arrow 5 marked cells 1 and unlabeled cells 2, which move evenly distributed in the laminar flow. Somewhat to the right of and below the microfluidic channel 4, a magnet 7 is arranged; the image immediately shows the accumulation of the marked cells on the bottom / substrate 8 of the channel within the magnetic field gradient 7. The GMR sensors, like all magnetoresistors, can also be connected to the Side walls of the channel wall and / or be arranged at the top of the channel. Again a little further to the right, ie in the direction of flow, are located at the bottom / substrate 8 of the channel several GMR sensors 3. By the "cell rolling" at the channel bottom and the enrichment of the marked cells by the external magnetic field, it is possible that as many as possible labeled cells can actually be detected by the GMR sensors.
Hier wird die Anreicherung von Zellen mit superparamagneti- schen Markern 1 aus einem komplexen Medium in einem Magnetfeld 9 gezeigt. Die laminare Strömung 5 verhindert eine Ver- wirbelung der Zellen 1 und 2. Durch Einstellung der Magnetfeldstärke können die Zellen 1,2 entlang der Substratoberfläche 8 rollen und kommen so in engsten Kontakt mit den GMR Sensoren 3. Die Stärke des Magnetfeldes soll aber den Transport der markierten Zellen im Mikrofluidikkanal nicht behin- dern, was beispielsweise durch einen geeigneten Pulsbetrieb sowie durch die Symmetrie des Gradientenfeldes eingestellt werden kann.Here, the enrichment of cells with superparamagnetic markers 1 from a complex medium in a magnetic field 9 is shown. The laminar flow 5 prevents swirling of the cells 1 and 2. By adjusting the magnetic field strength, the cells 1,2 can roll along the substrate surface 8 and thus come into closest contact with the GMR sensors 3. The strength of the magnetic field but the transport does not interfere with the labeled cells in the microfluidic channel, which can be adjusted, for example, by suitable pulsed operation and by the symmetry of the gradient field.
Rechts und im Abstand zu dem linken Teil der Figur 1 ist der Mikrofluidikkanal 4 im Querschnitt durch die Strömungsrichtung zu sehen. Zu erkennen sind die Feldlinien 9 des Magnetfeldes, die ihren Ursprung bei den GMR-Sensoren 3 haben und daher eine Gradientenverstärkung des Magnetfeldes bewirken. Dies ist maßgeblich darauf zurückzuführen, dass der Magnet 7 zu den GMR-Sensoren hin eine zumindest eine Schräge 6 hat, bevorzugt aber 2 Schrägen 6 wie gezeigt. Figur 2 zeigt das gleiche Bild wie Figur 1 im Längsquerschnitt und verdeutlicht das Cell-Rolling innerhalb der laminaren Strömung 5. Zu erkennen sind die drei Phasen des CeIl- Rollings, erstens (A) die Anreicherung der markierten Zellen 1 auf der Substratoberfläche des Bodens 8 des Mikrofluidikka- nals 4 im Magnetfeld 9, dann (B) das Cell-Rolling über die Sensoroberfläche wobei (C) die Zelldetektion stattfindet.On the right and at a distance from the left part of FIG. 1, the microfluidic channel 4 can be seen in cross-section through the flow direction. Evident are the field lines 9 of the magnetic field, which have their origin in the GMR sensors 3 and therefore cause a gradient amplification of the magnetic field. This is largely due to the fact that the magnet 7 has at least one bevel 6 towards the GMR sensors, but preferably 2 bevels 6 as shown. Figure 2 shows the same image as Figure 1 in longitudinal cross-section and illustrates the cell rolling within the laminar flow 5. The three phases of the CeIl- Rollings can be seen, first (A) the enrichment of the labeled cells 1 on the substrate surface of the bottom. 8 of the microfluidic channel 4 in the magnetic field 9, then (B) cell rolling over the sensor surface, where (C) cell detection takes place.
Nach einer vorteilhaften Ausführungsform der Erfindung wird, um eine Zelldetektion mit dem GMR Sensor (z.B. als Wheatsto- ne-Brückenschaltung) vorzunehmen, beispielsweise für eine kontinuierliche Anreicherung der Zellen das Gradienten- Magnetfeld (-100 mT mit dB/dx von einigen 10-100 T/m; abhängig von der Beladung der Zellen mit superparamagnetischen Partikeln) gepulst. Die Detektion der markierten Zellen erfolgt in einem schwachen Mess-Magnetfeld von ~ ImT.According to an advantageous embodiment of the invention, in order to perform a cell detection with the GMR sensor (eg as a Wheatstone bridge circuit), for example for a continuous enrichment of the cells, the gradient magnetic field (-100 mT with dB / dx of a few 10-100 T / m, depending on the loading of the cells with superparamagnetic particles) pulsed. The detection of the labeled cells takes place in a weak measuring magnetic field of ~ ImT.
In Figur 3 ist in zeitlicher Abfolge die Stärke des Magnetfeldes für die Zellanreicherung, Zelldetektion und die GMR- Messung gezeigt. Auf der X-Achse ist die Zeit aufgetragen, so dass zu erkennen ist, dass immer zwei Magnetfeldstärken im zeitlichen Wechsel angelegt werden. So kann ein Verfahren zur kontinuierlichen Zellanreicherung und Zelldetektion durch eine Abfolge von gepulsten Magnetfeldern durchgeführt werden.FIG. 3 shows in time sequence the strength of the magnetic field for cell enrichment, cell detection and GMR measurement. The time is plotted on the X-axis, so that it can be seen that always two magnetic field strengths are applied in temporal change. Thus, a method of continuous cell enrichment and cell detection may be performed by a sequence of pulsed magnetic fields.
Zu erkennen ist in Figur 3 die zyklische Abfolge von (1) Anreicherung, (2) +(3) Messung für eine kontinuierliche Messung, die bildlich graphisch dargestellt ist. Die Messung und Anreicherung der Zellen kann damit unabhängig voneinander im kHz Bereich verfolgt bzw. gesteuert werden. Zu erkennen ist in Figur 3 wie ganz oben mit einem „starken Magnetfeld und langen Pulszeiten" Zellanreicherung innerhalb des Mikroflui- dikkanals stattfindet. Darunter ist eine Graphik, die zeigt, dass ein schwächeres Magnetfeld mit geringerer Pulszeit zur Zelldetektion eingesetzt wird. Schließlich zeigt die unterste Graphik wie mit schwachem Magnetfeld und kurzer Pulszeit die GMR-Messung vollzogen wird. In Figur 4 ist wieder ein Mikrofluidikkanal zu sehen, wieder im Querschnitt senkrecht zur Strömungsrichtung wie in Figur 1 auf der rechten Seite.3 shows the cyclical sequence of (1) enrichment, (2) + (3) measurement for a continuous measurement, which is illustrated graphically. The measurement and accumulation of the cells can thus be tracked or controlled independently of each other in the kHz range. It can be seen in Figure 3 how at the very top with a "strong magnetic field and long pulse times" cell enrichment takes place within the microfluidic channel, below which is a graph showing that a weaker magnetic field with less pulse time is used for cell detection Graphic as with low magnetic field and short pulse time the GMR measurement is completed. FIG. 4 again shows a microfluidic channel, again in cross section perpendicular to the flow direction as in FIG. 1 on the right side.
Für die GMR Messung kann das Mess-Magnetfeld senkrecht oder in gleicher Ebene zu den GMR Sensoren angelegt werden (Figur 4) . Dabei kann der Magnet (Magnetjoch) des Gradientenmagnetfeldes zur Einstellung eines Gradienten im Mess-Magnetfeld benutzt werden, um eine lokale Verstimmung der Brückenglieder der GMR Messbrücke zu erreichen. Diese Verstimmung stellt das Messsignal für die Konzentration der magnetischen Partikel im Sensorbereich dar. In einer möglichen Ausgestaltungsform kann das Mess-Magnetfeld zusätzlich noch zeitlich moduliert werden, um z.B. mittels Lock-in Technik messen zu kön- nen und die niederfrequenten Rauschanteile (1/f- Rauschen) zu unterdrücken, um das Signal-Rauschverhältnis zu verbessern.For the GMR measurement, the measuring magnetic field can be applied vertically or in the same plane as the GMR sensors (FIG. 4). In this case, the magnet (magnetic yoke) of the gradient magnetic field can be used to set a gradient in the measuring magnetic field in order to achieve a local detuning of the bridge members of the GMR measuring bridge. This detuning represents the measurement signal for the concentration of the magnetic particles in the sensor region. In one possible embodiment, the measuring magnetic field can additionally be time-modulated in order, for example, to obtain a time delay. using lock-in technology and suppressing the low-frequency noise (1 / f noise) to improve the signal-to-noise ratio.
Nach einer vorteilhaften Ausführungsform wird mit gepulsten Magnetfeldern angereichert und detektiert wie in Figur 3 ge- zeigt. In Figur 4 wird dabei die schematische Anordnung der Magnete oder Spulen 7, 10 und 11 zur Anreicherung und Detek- tion um den Mikrofluidikkanal 4 gezeigt. Dabei wird beispielsweise der Magnet 7 für das starke Magnetfeld zur Anreicherung unterhalb der GMR-Sensoren angelegt und die Spulen 10 und 11 für das schwache Magnetfeld zur Detektion senkrecht zum GMR Sensor angelegt. Beide Felder können getrennt mit 2 Magneten gesteuert werden, wobei bevorzugt das schwache Magnetfeld in der Sensorebene angelegt wird.According to an advantageous embodiment, pulsed magnetic fields are enriched and detected as shown in FIG. In FIG. 4, the schematic arrangement of the magnets or coils 7, 10 and 11 for enrichment and detection around the microfluidic channel 4 is shown. In this case, for example, the magnet 7 for the strong magnetic field is applied for enrichment below the GMR sensors and the coils 10 and 11 for the weak magnetic field are applied perpendicular to the GMR sensor for detection. Both fields can be controlled separately with 2 magnets, wherein preferably the weak magnetic field is applied in the sensor plane.
Die wesentlichen Vorteile der erfindungsgemäßen Vorrichtung und des erfindungsgemäßen Verfahrens sind wie folgt:The essential advantages of the device according to the invention and of the method according to the invention are as follows:
1) Kontinuierliches Messverfahren um magnetisch markierte Zellen anzureichern und im Durchfluss zu erfassen.1) Continuous measurement method to enrich for magnetically labeled cells and to record in the flow.
2) Die Anreicherung der Zellen bzw. die ausgeübte Scherkraft auf die Zellen kann durch die Magnetfeldstärke und die Durchflussgeschwindigkeit gesteuert werden. 3) Markierte Zellen sind oberflächennah und können mit magnetoresistiven Bauteile sensitiv detektiert werden.2) The accumulation of the cells or the applied shear force on the cells can be controlled by the magnetic field strength and the flow rate. 3) Marked cells are near the surface and can be sensitively detected with magnetoresistive components.
4) Das vorgestellte Verfahren erlaubt eine großflächige Anwendung für Multiplexing (z.B. Array von GMR Sensoren) .4) The presented method allows a large area application for multiplexing (e.g., array of GMR sensors).
5) Das „Cell-Rolling" kann mit Hilfe von oberflächenfunkti- onalisierten Mikrofluidikkanälen an die Anwendung ange- passt werden. Die Funktionalisierung kann beispielsweise mit Rezeptoren (Selektinen) , biologische Komponenten (Proteinen, Polysaccharide), durch SAMs (self-assembled mono- layer) oder durch Silanisierung durchgeführt werden.5) Cell rolling can be adapted to the application by means of surface-functionalized microfluidic channels, for example functionalized with receptors (selectins), biological components (proteins, polysaccharides), or SAMs (self-assembled mono- layer) or by silanization.
6) Eine Anreicherung von markierten Zellen, wie seltene6) Enrichment of labeled cells, such as rare
Krebszellen (CTC; circulating tumor cells) , Tumorstammzellen, Inflammationszellen, Stammzellen, Bakterien oder Hefen, kann der eigentlichen Detektion im strömenden Medium vorausgehen .Circulating tumor cells (CTC), tumor stem cells, inflammatory cells, stem cells, bacteria or yeasts may precede the actual detection in the flowing medium.
7) Die magnetische Detektion kann mit optischen Methoden7) The magnetic detection can be done with optical methods
(FACS, Fluoreszenz, Absorption), und elektrischen Methoden (Impedanz, Dielektrophorese) kombiniert werden.(FACS, fluorescence, absorption), and electrical methods (impedance, dielectrophoresis) are combined.
8) Anwendungsfelder im Humanbereich sind unter anderem:8) Areas of application in the human area include:
Onkologie, regenerative Medizin, Infektiologie, klinische Diagnostik, klinische Chemie, Imaging. Oncology, regenerative medicine, infectiology, clinical diagnostics, clinical chemistry, imaging.

Claims

Patentansprüche claims
1. Vorrichtung zur Anreicherung und Detektion/Erfassung von Zellen, wobei in einem Kanal (4), in dem eine lamina- re Strömung (5) eines Mediums mit magnetisch markierten1. A device for enrichment and detection / detection of cells, wherein in a channel (4), in which a laminar flow (5) of a medium with magnetically marked
Zellen (1) fließt, zumindest ein Magnetowiderstand (3) in zumindest einem äußeren, ihn umgebenden Magnetfeld (9) angeordnet ist.Cells (1) flows, at least one magnetoresistor (3) in at least one outer, surrounding magnetic field (9) is arranged.
2. Vorrichtung nach Anspruch 1, bei der das Medium ein komplexes Medium wie Blut ist.2. Device according to claim 1, wherein the medium is a complex medium such as blood.
3. Vorrichtung nach Anspruch 1 oder 2, wobei der Ursprung der Feldlinien des Magnetfeldes (9) bei dem zumindest ei- nen Magnetowiderstand (3) liegt.3. Apparatus according to claim 1 or 2, wherein the origin of the field lines of the magnetic field (9) in the at least one magnetoresistor (3).
4. Vorrichtung nach einem der vorstehenden Ansprüche, wobei der Kanal (4) ein Mikrofluidikkanal ist.4. Device according to one of the preceding claims, wherein the channel (4) is a microfluidic channel.
5. Vorrichtung nach einem der vorstehenden Ansprüche, wobei zur Erzeugung des umgebenden Magnetfeldes zumindest ein Magnet (7) unterhalb des zumindest einen Magnetowiderstandes (3) angeordnet ist.5. Device according to one of the preceding claims, wherein for generating the surrounding magnetic field at least one magnet (7) below the at least one magnetoresistor (3) is arranged.
6. Vorrichtung nach einem der vorstehenden Ansprüche, wobei der Magnetowiderstand (3) ein GMR-Sensor ist.6. Device according to one of the preceding claims, wherein the magnetoresistor (3) is a GMR sensor.
7. Vorrichtung nach einem der vorstehenden Ansprüche, wobei ein weiteres Magnetfeld vorgesehen ist.7. Device according to one of the preceding claims, wherein a further magnetic field is provided.
8. Vorrichtung nach einem der vorstehenden Ansprüche, wobei ein weiteres Magnetfeld durch eine in unmittelbarer Nähe zum Magnetowiderstand angeordnete Spule (10,11) erzeugt wird.8. Device according to one of the preceding claims, wherein a further magnetic field by a arranged in close proximity to the magnetoresistor coil (10,11) is generated.
9. Vorrichtung nach einem der vorstehenden Ansprüche, wobei eine Spule (10,11) senkrecht zu dem Magnetowiderstand angeordnet ist. 9. Device according to one of the preceding claims, wherein a coil (10,11) is arranged perpendicular to the magnetoresistor.
10. Verfahren zur Anreicherung und Detektion von magnetisch markierten Zellen in einem laminar strömenden Medium, wobei Zellen an einem Magnetowiderstand durch zumindest ein äußeres Magnetfeld angereichert werden.10. A method for enrichment and detection of magnetically labeled cells in a laminar flowing medium, wherein cells are enriched in a magnetoresistor by at least one external magnetic field.
11. Verfahren nach Anspruch 8, wobei der zumindest eine Magnetowiderstand im Kanal in gepulstem Betrieb von zumindest zwei Magnetfeldern umgeben ist.11. The method of claim 8, wherein the at least one magnetoresistance in the channel is surrounded in pulsed operation of at least two magnetic fields.
12. Verfahren nach Anspruch 9, wobei die zumindest zwei Magnetfelder, die gepulst angelegt werden, sich in Größenordnungen an Stärke unterscheiden.The method of claim 9, wherein the at least two magnetic fields applied in a pulsed manner differ in magnitudes in magnitude.
13. Verfahren nach Anspruch 8 oder 9, wobei die zumindest zwei Magnetfelder sich in ihrer Stärke um zumindest einen Faktor 100 unterscheiden. 13. The method of claim 8 or 9, wherein the at least two magnetic fields differ in strength by at least a factor of 100.
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US20110315635A1 (en) 2011-12-29

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