WO2011104312A1

WO2011104312A1 - Improvement of the detection limit of magnetically labelled samples

Info

Publication number: WO2011104312A1
Application number: PCT/EP2011/052767
Authority: WO
Inventors: Franz Cemic; Jürgen Hemberger
Original assignee: Fachhochschule Giessen-Friedberg
Priority date: 2010-02-24
Filing date: 2011-02-24
Publication date: 2011-09-01
Also published as: DE102010009161A1

Abstract

The invention describes an apparatus and a method for reducing the detection limit for magnetically labelled molecules. This is achieved by way of screening the sensor against the Earth's magnetic field and the electronics. Furthermore, the sample slide is cleaned using a magnet before the measurement and calibrated for its own magnetization, such that these values can be included in a calibration with a sample after measurement.

Description

Patent application

TITLE

Improvement of the detection limit of magnetically labeled samples

The present invention relates to an apparatus and method for increasing the detection limit of devices that characterize and quantify magnetically-labeled samples. [Description and Introduction of the General Field of the Invention]

The invention relates to increasing the sensitivity of devices for measuring magnetically labeled samples.

[State of the art]

In biological analysis technology increasingly magnetizable nanoparticles or microparticles (beads) are used on carriers (biochips) for the detection of molecules. For this purpose, magnetic fields are applied, with which the beads are magnetized. The resulting magnetic field is measured. Measurand is the strength of the resulting magnetic fields.

DE60214674 describes a sensor for measuring magnetic particle accumulations, the measurement being independent of the magnetic background. The document explicitly stated that the magnetic background should not play a role for the measurement with a Hall sensor. A similar sensor is described in US 6046585.

In DE 102006055957 it is shown that the stray field is strong enough only near the surface of the bead in order to be detected. Already at a distance of about 50 μιτι the signal decreases so quickly that a bead with XMR sensors (eg giant magneto resistance (GMR), tunnel magneto resistance (TMR), anisotropic magnetic resistance (AMR), colossal magnetic resistance ( CMR), organic magnetic resistance (OMR)) can no longer be detected with sufficient sensitivity [0042]. In addition, only 2/3 of the surface is used for detection by an XMR sensor. The remainder contributes to the noise in [0046].

A special magnetic shielding of the excitation coil relative to the sensor coil is mentioned in document DE 10137665 [0032].

The prior art assumes that shielding the magnetic background is not necessary.

This has the disadvantage that the sensitivity for measuring magnetic particles on membrane strips is severely limited.

In the devices currently produced, the magnetization of the sample carrier is not measured. This has the disadvantage that a calibration of the measured value with respect to the residual magnetization on the lateral flow assay (LFA) strip is not possible. There is thus too much background noise to measure the magnetism of the LFA strip (carrier) and the magnetism of minute magnetic particles. [Task]

Object of the present invention is to eliminate the disadvantages of the prior art by means of the described arrangement.

[Solution of the task]

This object is achieved by reducing the magnetic noise floor and an additional correction of the magnetic background. To reduce the magnetic noise floor, an electromagnet with a field strength in the range of 100 to 10000 μΑ / m ² , more preferably in a range of 10000 to 100000 μΑ / m ² is connected in front of the measuring device. The electromagnet is separated from the measuring device by a magnetic shield.

In addition, all parts of the measuring device (sensor, sample carrier, in particular the electronics of the device) are shielded from internal and external interference fields. Surprisingly, this results in a reduction of the magnetic field strength in the sensor interior to up to 1/100 of the local earth magnetic field (eg reduction of the magnetic flux density from 3.1 10 ^"5 T to approximately 3.1 10 ^{" 7} T; for comparison, magnetic flux densities in space: 10 ^"10 to 10 ^{" 8} T). This has the advantage that the shielding and cleaning makes it possible to measure significantly smaller magnetic flux densities in the sensor interior and thus to increase the sensitivity of the measurement. The magnetic shield also includes the entire meter. Separate magnetic shields for the device, the electronics, the electromagnet and the measuring device can thus be installed.

The magnetic shield comprises several devices. These include plastic films, in which preferably soft magnetic particles are incorporated. It can also be used metal foils made of hard or soft magnetic metals or alloys.

Similarly, metal plates with 1 μιτι be used up to 10 mm thickness.

Sheets, foils, particles, metals or alloys include, by way of example, the following shielding materials, such as:

Alloys of iron with nickel, cobalt e.g. Mu metal (nickel-iron alloy (75-81% Ni, 0-5% Co), Permenorm (about 50% Ni, balance iron), Vacoflux (about 49% Co, 2% V, balance iron);

Alloys with iron with silicon e.g. 85% iron, 9% Si, 6% Al) or (15 to 38% silicon or boron; 0 to 10% Cr, Nb, Ti, V, Ta, Mo, W, Mn, Co,

Ni, Ba Sr; Rest of iron)

Copper, aluminum, nickel, cobalt, lead, iron, iron nitride or ferrites, eg hematite (Fe ₂ O ₃ ), maghemite (Y-Fe ₂ O ₃ ) or magnetite (Fe ₃ O ₄ ), also in combination with Ni, Zn, Mn

and composites e.g. Iron - copper; Iron - aluminum; Iron - silver; The aforementioned materials are used with or without earthing.

This magnetic shielding measures a very small number of magnetic particles in a spatially resolved manner. The sensor used is preferably Hall sensors or fluxgate sensors. However, XMR sensors or Superconducting Quantum Interference Device (SQUID) sensors can also be used.

A sample carrier, preferably a membrane carrier typical of lateral flow assays, is inserted into the device. This sample carrier generally consists of a carrier plate 101 and a suitable membrane to which substances 103 to 107 are applied.

In general, the sample carrier still has a residual ferromagnetism (ferromagnetic or paramagnetic particles of separating tools) due to its production, which clearly influences the measuring accuracy. Therefore, the sample carrier without sample is passed one or more times through a cleaning magnet (for example, a permanent magnet, but preferably an electromagnet, more preferably a pole-changing electromagnet) 201 and thus cleaned of these particles.

The cleaning magnet likewise comprises a tube 402 surrounded by a coil (for example made of copper) 401 with superparamagnetic or ferromagnetic particles or nanoparticles. A particularly preferred arrangement is the use of superparamagnetic nanoparticles, since they have the advantage of having no magnetic hysteresis.

The cleaning magnet may be surrounded by a two-part magnetic shield 403 and 404, wherein the lower shield 404 is designed as a shell and displaceable.

The process of cleaning ferro- or paramagnetic particles can be carried out with a single pull through the sample carrier. Preferably, however, first a magnetic measurement of the sample carrier is carried out. Subsequently, the cleaning of ferro- or paramagnetic particles takes place by performing the sample carrier under the cleaning magnet. Subsequently, a magnetic measurement is performed again. If there are any deviations from the previous measurement, the steps will continue as long

- Carrying out the sample carrier under the cleaning magnet and

- Carrying out the magnetic measurement

repeated until no deviations occur.

There are no deviations if the measured differences are within a range of 10 ^"2 T to 10 ^{" 9} T, preferably 3.1 10 ^"5 T to 3.1 10 ^{" 7} T. After this cleaning process is completed, the solenoid 201 is moved into a shielded area of the meter. Thereafter, the sample carrier is calibrated without a sample. For this purpose, with a shielded sensor 202 according to the invention, the magnetism of the sample carrier is measured and stored as a function of the position (eg length LFA strips). These values can be determined and stored at individual locations or over the entire length.

Subsequently, the sample carrier is given the sample, by the application of which the superparamagnetic molecule-particle conjugates are released from the conjugate pad. In this case, conjugated molecules include biomolecules such as proteins, in particular antibodies and / or antigens, nucleic acids, in particular DNA, RNA, LNA (locked nucleic acid, DNA ribose: C2 and C4 are linked via an oxygen-methylene bridge), PNA (peptide-protein) Nucleic acid; DNA: base

Bose phosphate backbone - PNA: peptide backbone

TNA

(Threose - Nucleic Acid; DNA: Ribose Phosphate Backbone - TNA: threose phosphorodiester backbone), GNA (glycol nucleic acid, DNA: Ribose Phosphate Backbone - GNA: Glycerol Phosphordiester Backbone) etc., as well as their fragments, but also be chemical substances that act as antigens.

Subsequently, the magnetism of the sample carrier loaded with the sample is measured and recorded as a function of the position. The measured value, ie the magnetization as a function of the location on the test strip provided with the sample, is corrected with the previously determined and stored background values.

According to the invention, very small amounts of beads in the range of a few (1 to 100) or (10 to 1000) pg per test line of the superparamagnetic molecule-particle conjugates described above can be detected by this method.

[Embodiments]

In Fig. 1, a sample carrier is shown. This consists of a support plate 101 made of plastic. Reagentienpads (membranes, tissue, cellulose) 103, 104, 105 are attached to the membrane 107. Päd 103 is used for sample collection, 104 contains the superparamagnetic molecule-particle conjugates. The sample flows from the pedicle 103 to the pad 104, with the analyte binding specifically to the molecule-particle conjugates. During flow along the membrane 107, individual sample components are held in the regions 106. The remaining components of the sample are collected in the membrane 105.

The measuring cycle begins with the cleaning of the sample holder without a sample. For this purpose, the electromagnet 201 is turned on and the sample carrier is guided 1 to 3 times by the electromagnet, wherein the electromagnet is turned off during the return movement in each case. After the cleaning procedure, the turned-off solenoid 201 is shifted to a shielded area. Alternatively, a shield in front of the electromagnet can be performed. Residual magnetic fields are intercepted by the magnetic shield 205 of the measuring device.

Subsequently, the magnetization of the cleaned sample carrier is detected without a sample. To do this, the magnetizations are sensed by the sensor 108, e.g. Hall sensor, fluxgate, XMR sensor measured and stored as a function of the position over the entire length 110. If only certain collection areas 106 are provided, it is only necessary to measure and store the magnetization for individual lengths 109.

Subsequently, the sample is placed on the sample carrier. After the sample has reached the membrane 105, the magnetization is again registered position-dependent over the length 110 or at individual lengths 109. The measured values of the sample are corrected with the previously determined values of the cleaned test strip and a distribution of the magnetization as a function of the location on the membrane 107 is obtained. 2, a measuring device according to the invention is shown schematically. The measuring device 202 is separated from the electronics 203 and the electromagnet 201 by a magnetic shield 204 and 205. The electronics are connected to the measuring device by a shielded cable 206. In the case of the electric magnet 201, the magnetic shield 205 is preferably designed to be movable (positions A and B). When the magnetic shield 205 is in position B, the solenoid is released and the sample carrier 207 can be cleaned. The ferromagnetic or paramagnetic particles are thereby removed by the electromagnet 201 from the sample carrier 207. In order to facilitate cleaning of the electromagnet, the device 208 captures the ferromagnetic or paramagnetic particles. The device 208 is made of plastic, paper (eg filter paper) or textiles (eg typewriter belts with liquid - better adhesion of the particles by moisture). So that the sample carrier 207 is not contaminated again, first the magnetic shield 205 is brought into position A. Thereafter, the electromagnet is switched off.

Alternatively, the device 208 is to be replaced after guiding the magnetic shield 205 in position A and before switching off the electromagnet 201. This can be realized in a textile tape as a winding and rewinding of the tape.

The shield 205 is used as a receptacle, e.g. Shell executable.

After the electromagnet 201 has been switched off, the calibration of the sample carrier 207 takes place. Subsequently, the sample carrier is measured with the sample, the data of the calibration being used to correct the measured data. In FIG. 3, the measuring device 302 is also the electromagnet at the same time. The measuring device here also has a magnetic shield 304 in relation to the electronics 303. The control of the measuring device or the electromagnet via the cable 306. In the case of the circuit as an electromagnet, the interception devices 308 (corresponding to the devices 208) are introduced. Once the sample carrier 307 has been cleaned, the interceptors 308 are guided behind the magnetic shield 304.

[Illustration legends and reference list]

1 is a schematic view of the sample carrier (for example, lateral flow assay)

Fig. 2 is a schematic view of the measuring device with additional cleaning magnet

Fig. 3 is a schematic view of the measuring device with measuring sensor, which can be switched as a cleaning magnet

Fig. 4 is a schematic view of an embodiment of a cleaning magnet

Claims

[Claims]

1 . A device for improving the detection of magnetically marked substances, characterized in that the device comprises at least one cleaning magnet.

2. A device according to claim 1, characterized in that the cleaning magnet comprises a permanent magnet, an electromagnet or a combination of the magnets.

3. A device according to any one of claims 1 to 2, characterized in that the cleaning magnet comprises a coil with a tube, wherein the tube has superparamagnetic nanoparticles.

4. A device according to one of claims 1 to 3, characterized in that the cleaning magnet comprises a device (208, 308) for trapping para or ferromagnetic substances.

5. A device according to one of claims 1 to 4, characterized in that the measuring device comprises the cleaning magnet.

6. A magnet according to any one of claims 1 to 5, characterized in that the magnet comprises a coil with a tube, wherein the tube has superparamagnetic nanoparticles.

7. A method for improving the detection of magnetically marked substances, characterized in that the sample carrier is cleaned without sample of adhering para or ferromagnetic substances magnetically.

8. A method according to claim 7, characterized in that

a) the sample carrier is measured magnetically without a sample,

b) the sample carrier is cleaned magnetically without a sample,

c) the sample carrier is measured magnetically without a sample,

d) repeating steps b) and c) are repeated until between two measure- ments deviations from 0 to 3 10 ^"5 T, preferably 1 ^{10" 7} T can be achieved.

9. A method according to any one of claims 7 to 8, characterized in that the measuring device is switchable as a cleaning magnet.