WO2006056001A1 - Dispositif de determination qualitative et quantitative d'analytes - Google Patents

Dispositif de determination qualitative et quantitative d'analytes Download PDF

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Publication number
WO2006056001A1
WO2006056001A1 PCT/AT2005/000477 AT2005000477W WO2006056001A1 WO 2006056001 A1 WO2006056001 A1 WO 2006056001A1 AT 2005000477 W AT2005000477 W AT 2005000477W WO 2006056001 A1 WO2006056001 A1 WO 2006056001A1
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WO
WIPO (PCT)
Prior art keywords
hollow body
distributor
particles
analyte
outlet opening
Prior art date
Application number
PCT/AT2005/000477
Other languages
German (de)
English (en)
Inventor
Thomas Schlederer
Original Assignee
Thomas Schlederer
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Application filed by Thomas Schlederer filed Critical Thomas Schlederer
Publication of WO2006056001A1 publication Critical patent/WO2006056001A1/fr

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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
    • B03C1/025—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
    • 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
    • B03C1/0332—Component parts; Auxiliary operations characterised by the magnetic circuit using permanent magnets
    • 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
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G01N35/10—Devices for transferring samples or any liquids to, in, or from, the analysis apparatus, e.g. suction devices, injection devices
    • G01N35/1095—Devices for transferring samples or any liquids to, in, or from, the analysis apparatus, e.g. suction devices, injection devices for supplying the samples to flow-through analysers
    • G01N35/1097—Devices for transferring samples or any liquids to, in, or from, the analysis apparatus, e.g. suction devices, injection devices for supplying the samples to flow-through analysers characterised by the valves
    • 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
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G01N35/0098—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor involving analyte bound to insoluble magnetic carrier, e.g. using magnetic separation

Definitions

  • the present invention relates to a device for the qualitative and quantitative determination of analytes.
  • NPT near patient testing
  • Areas where such procedures may be used include the areas of blood clotting, emergency diagnostics, markers of heart disease (heart attack), urinalysis, clinical chemistry, immunology, and metabolic disorders.
  • the US 6,133,043 relates to a device for analyzing a sample comprising a cell having a defined volume and an electrode which is capable of applying an electrical voltage to the sample, a magnetic Auffangvor ⁇ direction with one or more magnets, a voltage source for generating a Voltage at the electrode sufficient to induce luminescence of an electrochemiluminescent substance in the sample, and a light detector.
  • the device comprises inter alia a detection device coupled to a measuring cell, a distributor and a pump.
  • the distributor has a plurality of inlet openings and an outlet opening for distributing various solutions (including the sample mixture itself) into the tubular hollow bodies connected to the distributor, which are connected to the pump via the magnetic device and the detection device.
  • Such an arrangement leads to a high mechanical load of any magnetic particles used.
  • US 2003/0127396 an apparatus for mixing and separating magnetic particles is disclosed.
  • This device comprises, inter alia, a container which serves for receiving liquids and magnetic particles, and a magnetic device in the immediate vicinity of the container. In this container, one or more tubes can be inserted through an opening at the top, with the help of which washing liquids are transported in and out of the container.
  • WO 2004/011146 describes an apparatus for mixing, separating, collecting and washing magnetic particles, which includes, inter alia, a pipette whose upper end is arranged on a pressure device and a magnet.
  • WO 1994/11078 relates to a device for immobilizing and isolating cells or analytes bound to magnetic particles from a liquid medium.
  • This device comprises inter alia magnetic means for generating a magnetic field and tubular hollow bodies for transporting the liquid medium.
  • EP 0 687 501 B1 discloses a method and a device in which magnetic particles in pipettes can be temporarily immo-stabilized during an examination with the aid of a magnet.
  • the results of the NPT can be applied quickly and directly in the operating theater, in intensive care units, outpatient clinics, blood banks or directly in the doctor's office.
  • the present invention relates to a device for determining at least one analyte in a sample
  • a device for determining at least one analyte in a sample comprising a distributor having at least one inlet opening for introducing liquids, suspensions and / or gases into the distributor and having at least one outlet opening for removing liquids, suspensions and / or gases from the distributor in which a tubular hollow body connected to the distributor is attached to at least one outlet opening, to which a magnetic device for temporary immobilization of magnetic analyte-binding particles introduced into the interior of the hollow body through an inlet opening or inside the hollow body an inner boundary of the hollow body is arranged, wherein the distributor a displacer is arranged.
  • Liquids and / or gases refers to all those liquids that can be used in an analysis, which apart from the sample liquid are usually salt solutions, organic solutions, buffers, water, suspensions etc. or air, noble gases, inert Gases, etc., on the one hand serve to strengthen the binding analyte particles and clean the Vor ⁇ direction and on the other hand are able to displace any liquid from the device.
  • Tubeular hollow bodies are tubular hollow bodies which are open at at least two ends and are also used, for example, in chromatography .These as well as all those components which come into contact with the fluids used should be inert to the solutions used the tubular hollow body, for example, consist of metal, but appropriate plastic is preferred.
  • “Analytically binding particles” are magnetic particles which are capable of specifically binding an analyte from a sample.
  • the particles can have paramagnetic, superparamagnetic or ferromagnetic properties according to the invention, with paramagnetic and superparamagnetic particles preferably being used.
  • Magnetic particles can be immobilized by the application of a magnetic field on a surface and optionally concentrated, whereby the handling of such particles within the device is facilitated.
  • Particles which are particularly suitable for the use according to the invention are described, for example, in "Biomagnetic Techniques in Molecular Biotechnology”. logy ", Dynal, Technical Handbook 3 rd Edition ⁇ 1998.
  • an outlet opening can also serve to remove waste liquid from the device and to sufficiently flush the lines before using the device.
  • the inlet and outlet openings are preferably designed such that they are optionally closable, in order to prevent the escape of liquid or gas from the device during the changeover of the distributor when not using an opening.
  • the closing could be carried out, for example, by attaching a valve.
  • a device according to the invention has advantages over hitherto known devices for determining analytes to which magnetic particles are also used for binding the analytes to be examined.
  • the magnetic particles are temporarily immobilized in pipettes during the washing steps, the incubation of the particles with the sample, with a second label-conjugated analyte-binding partner and the subsequent detection reaction in containers.
  • this device has the disadvantage that a correspondingly large sample volume and many pipetting steps are required, for which new containers with the corresponding liquids (eg washing solutions) must be made available in each case.
  • small sample volumes can be carried out in one and the same hollow body.
  • a displacer is arranged on the distributor. This can be connected to the distributor via its own positive displacement inlet opening.
  • a displacer serves to introduce a liquid or a gas into the device or to remove.
  • the displacer is preferably attached to the distributor at an inlet opening (so-called displacement inlet opening) via a further tubular hollow body.
  • the displacer should be able to move the substances to be transported in the tubular hollow bodies both forward and backward in order, if appropriate, to permit mixing of the magnetic particles in the tubular hollow bodies which are arranged at an opening.
  • the arrangement of the displacer directly on the distributor via its own displacer input opening has a number of further advantages.
  • the displacer is designed as a pump or syringe.
  • pumps and syringes are particularly well suited for bringing liquids into the device according to the invention and for moving them within them, in particular commercially available syringes and pumps, which are also used in chromatography can.
  • the distributor is designed as a rotary or multi-way valve.
  • Rotary and multi-way valves are characterized by the many Due to the large number of openings, the distribution options are mainly due to their high degree of automation. Therefore, such valves are particularly well liquids and gases to distribute in the device.
  • the distributor comprises a mixing chamber.
  • the mixing chamber has a stirring device.
  • a stirring device This substantially facilitates the generation of a homogeneous mixture in or at the distributor.
  • Such mixing devices are already known for use in chromatography and can thus be used directly for the purpose of the invention. Nevertheless, it is entirely possible to dispense with a stirring device when the mixing of the liquids takes place by means of flow effects.
  • Tumbular hollow bodies are preferably attached to the at least one inlet opening.
  • tubular hollow bodies serve to direct liquids and gases over greater distances into the distributor, whereby e.g. Hoses commonly used in chromatography are particularly well suited.
  • the tubular hollow body is designed as Kapilla ⁇ re.
  • the use of capillaries makes it possible to use small volumes of sample and liquid, on the one hand small amounts of sample sufficient to carry out investigations, and on the other hand miniaturization of the device according to the invention is made possible.
  • the inner diameter of the capillaries used is a maximum of 3 mm, preferably a maximum of 2 mm, maximum lmm, maximum 0.5mm, maximum 0.3mm, maximum 0.2mm, maximum 0.1mm, maximum 500 ⁇ m.
  • the minimum inner diameter depends on the size or diameter of the functionalized magnetic particles used, the inner diameter should be at least twice or at least three times as large as the diameter of the particles.
  • the hollow body connected to the outlet opening has at least one cross-sectional widening, in particular at least one chamber.
  • a cross-sectional widening of the tubular hollow body serves, for example, to form a kind of chamber, in which the mixing of the particles with solutions introduced into the device is promoted due to the currents generated by the flow and the resulting turbulences.
  • the chambers may optionally be delimited on one side with a liquid-permeable barrier.
  • a tubular hollow body attached to an outlet opening has a plurality of cross-sectional enlargements occurring one behind the other, the hollow body being one or more pieces, i. a plurality of hollow bodies are connected to each other, is formed.
  • a multi-part arrangement of several cross-sectional enlargements would make it possible, for example, to provide different chambers with differently functionalized particles and thus allow the analysis of several analytes simultaneously.
  • a plurality of outlet openings may alternatively be provided with tubular hollow bodies according to the invention. As a result, several hollow bodies can be addressed individually by the changeover of the distributor.
  • the tubular hollow body connected to the outlet opening has at least partially transparent regions.
  • Transparent areas are particularly advantageous if an event, such as a color reaction, should be made visible within the hollow body. Also, a position control of the particles located in the hollow body would be possible with appropriate particle size or density. These ranges should, if, for example, photometric measurements are carried out directly on the hollow bodies, the light should pass through as unhindered as possible. to let. Therefore, the transparent areas may also consist of materials other than the hollow body (eg glass).
  • the transparent regions are preferably designed as measuring cells.
  • Measuring cells which can be used according to the invention are already known in photometry, chromatography, etc. as flow cells and can be introduced into the flow of the device accordingly.
  • the means for immobolization is a magnetic device which is arranged to be relatively movable with respect to the hollow body connected to the outlet opening of the distributor.
  • the analyte-binding particles which are preferably of super, para- or ferromagnetic nature
  • a magnetic device is arranged movably, whereby the area of the immobilization can be flexibly selected.
  • the magnetic device is arranged to be movable at least approximately perpendicularly to the tubular hollow body. Furthermore, such an arrangement allows the use of a permanent magnet which can be moved away from the hollow body when the magnetic field is no longer desired on the hollow body.
  • the magnetic device comprises an electric or permanent magnet.
  • An electromagnet has the advantage that the magnetic field can be controlled by means of a circuit. Nevertheless, permanent magnets are also particularly suitable for use in the device according to the invention.
  • a filter or a filter combination is attached to at least one inlet opening or to at least one outlet opening or to at least one tubular hollow body connected thereto.
  • filters at the inlet or outlet openings or at the tubular hollow bodies connected thereto has a number of advantages.
  • solid substances can be removed from the sample before they enter the device and interfere with the analysis.
  • Fer ⁇ ner can be removed by appropriate filters and other unwanted sample components (eg leukocytes in whole blood) become.
  • Other substances which interfere with the analysis can also be removed from the sample by the use of substance-specific filters (compare affinity column in chromatography).
  • the sample preparation can be omitted partially or completely, whereby the use of the Vor ⁇ direction of the invention, for example, in emergency medicine is particularly advantageous.
  • the filter is a plasmapheresis filter.
  • filters for the immune platform are those whose structure is suitable for separating cell components from whole blood. Such filters are used in blood purification for dialysis machines or in lateral flow assays (for example from Schlei cher & Schuell, "AccuFlow Plus").
  • a detector is arranged on the hollow body connected to the outlet opening of the distributor, the detector being selected from the group consisting of a photometer, fluorometer, scintillation detector, CCD detector and combinations thereof.
  • the arrangement of a detector on the hollow body, which is attached to the Aus ⁇ lassö réelle, has proved to be particularly advantageous, since it is thereby made possible to analyze the binding of an analyte of particles directly in the hollow body. It is essential in such an embodiment that the signal to be measured can penetrate the hollow body at the position of the detector.
  • the hollow body is transparent when a color reaction or fluorescence is measured.
  • the detector can also be positioned outside the device according to the invention or not on the hollow body. In this case, the analyte bound to the particles is transferred as a suspension into further vessels and subsequently determined in a separate detector.
  • the device according to the invention is designed as a cassette.
  • the design of the device as a cassette makes it possible to flexibly combine the device with other other analyzers (eg blood gas machines).
  • the cassette according to the invention can consist only of tubular hollow bodies (in which optionally already functionalized magnetic particles are present), which are introduced into a device which corresponds to the invention a distributor, a displacer and at least one magne ⁇ tables device provides.
  • a distributor, a displacer and at least one magne ⁇ tables device provides.
  • the cassette can be introduced by a few simple steps.
  • the cassette may additionally comprise a distributor or a displacer or a magnetic device or combinations thereof.
  • a further aspect of the present invention relates to a method for determining at least one analyte in a sample with a device according to the invention comprising the steps:
  • sample-particle suspension optionally introducing the sample-particle suspension into the hollow body connected to the outlet opening of the distributor, if the incubation was not carried out in the hollow body,
  • the magnetic particles can be immobilized on the hollow body during the entire process or only for example when changing the solutions or when washing the particles.
  • the detection takes place inside or outside the hollow body connected to the outlet opening of the distributor.
  • the detection of analytes bound to magnetic particles can be carried out as described above with a wide variety of detectors, depending on which analyte is to be detected by which detection means.
  • the detection can take place either directly in the hollow body or after the particles loaded with analyte from this hollow body into a vessel (eg in a cuvette for a photometer or a microtiter plate) was transferred.
  • the analyte For further characterization of the analyte, it may be separated from the particles and further analyzed, e.g. Mass spectrometry. This separation can take place, for example, by changing the pH conditions, salt concentration or by addition of a competitive substance displacing the analyte (for example analogs) (see chromatographic methods).
  • the sample is selected from the group consisting of body fluids, in particular blood, blood serum and saliva, urine, perspiration, tissue samples, cell culture supernatants, in particular eukaryotic and prokaryotic culture supernatants, environmental analytical solutions and puncture liquids of various organs.
  • body fluids in particular blood, blood serum and saliva, urine, perspiration, tissue samples, cell culture supernatants, in particular eukaryotic and prokaryotic culture supernatants, environmental analytical solutions and puncture liquids of various organs.
  • Extraction or separation can be carried out by suitable solvents or by homogenization and removal of the solid constituents (by centrifugation or filtration), it being possible for filtration to be carried out directly on the apparatus according to the invention or within it (eg by providing Filtering on the device according to the invention).
  • the magnetic analyte-binding particles have at least one chemical modification.
  • the particles By chemically modifying the particles, they can be functionalized so that the particles are able to specifically bind an analyte from a sample.
  • the chemical mixed modification an analyte-specific binding partner.
  • the chemical modification comprises a substance selected from the group consisting of proteins, in particular antibodies, antibody fragments, protein A, protein G and lectins, biotin, streptavidin, avidin, phages, phage proteins, viruses, vi ⁇ rale proteins / particles , Nucleic Acids, Oligonucleotides, and combinations thereof (see “Antibodies, a laboratory manual”, Ed Harlow, David Lane; ColD Spring Harbor Laboratory Press, 1988).
  • the means for detecting the analyte comprises a further binding partner of the analyte.
  • the use of a corresponding binding partner is particularly well suited, which if appropriate is more easily or to a greater extent detectable than the analyte to be determined.
  • the further binding partner is conjugated with a marker.
  • the binding partner which binds to the analyte on the particles, may be conjugated to a label.
  • markers are e.g. from immunology (see ELISA, RIA and the like). Above all, a marker serves to indicate the presence of a conjugate directly or indirectly via a further reaction.
  • the marker is an enzyme, in particular horseradish peroxidase, ⁇ -galactosidase and alkaline phosphatase, radioactive isotope, BrDU, digitonin, Ki67, PCNA, ruthenium compounds, such as Ru chelates, tris (2,2'-bipyridines) ruthenium (II) (see, for example, WO92 / 14138), or combina tions thereof.
  • an enzyme in particular horseradish peroxidase, ⁇ -galactosidase and alkaline phosphatase, radioactive isotope, BrDU, digitonin, Ki67, PCNA, ruthenium compounds, such as Ru chelates, tris (2,2'-bipyridines) ruthenium (II) (see, for example, WO92 / 14138), or combina tions thereof.
  • the means for detection preferably comprise a substrate, which is reacted by the label of the conjugated binding partner, which substrate can be selected from the group consisting of luminol, dixethanes, acridine esters and combinations thereof.
  • an analyte on the particles can be easily determined by methods such as PCR, ELISA or PAP-APAAP.
  • the volume of the sample is preferably from 1 ⁇ l to 1 ml, preferably from 2 ⁇ l to 500 ⁇ l, in particular from 5 ⁇ l to 200 ⁇ l.
  • any sample volumes can be analyzed with the device disclosed herein, but preferably small volumes are used in certain fields of application (for example a blood sample in a premature infant in the clinic).
  • small volumes up to a maximum of 5 ml, a maximum of 4 ml, a maximum of 2 ml, a maximum of ImI, a maximum of 500 ⁇ l, a maximum of 200 ⁇ l in the method according to the invention is made possible above all by the provision of a corresponding device.
  • the device according to the invention is used in combination with a Blut ⁇ gas automaton, whereby the use of a blood gas machine with respect to the determination of analytes in the blood can be dramatically expanded.
  • the device according to the present invention can be connected in a simple manner with the aid of capillaries, connecting pieces and valves to an outlet opening or output capillary of a blood gas automat.
  • the device can also be designed as a disposable or reusable cassette, which can be exchangeably introduced into a blood gas automat.
  • Automatic blood gas analyzers are sufficiently well known to the person skilled in the art, for example, by the companies Roche (“Roche COMPACT 3") and Osmetech ("Osmetech OPTI CCA Blood Gas Analyzer").
  • FIG. 1 shows an apparatus for determining an analyte which comprises a distributor 1 which has a plurality of inlet openings 2-6, an inlet opening 7 for mounting a displacer 8 (FIG. drängereingangsö réelle) and an outlet opening 9 has.
  • a tubular hollow body 10 is mounted, on which a magnetic device 11 is arranged.
  • a detector 12 for direct or indirect determination of the analyte immobilized on the magnetic particles is likewise arranged along the hollow body 10.
  • the sample and other required reagents such as buffers and the like, sucked and optionally in ⁇ with the magnetic particles in the device according to the invention in ⁇ cubiert.
  • the resulting suspension can be mixed in a mixing chamber.
  • this suspension can be pumped into the tubular hollow body 10 of the outlet and mixed with or without forward and backward movement of the suspension.
  • the analyte to be determined is bound by appropriate interactions to the magnetic particles suspended in the solution.
  • the magnetic particles are retained in the outlet hollow body 10 by applying a magnetic field to its outer boundary within the boundary.
  • the entire volume of the suspension must be guided past the applied magnetic field by means of a displacement 8 by means of a corresponding displacement and return movement in order to quantitatively immobilize the magnetic particles.
  • a washing process is then carried out by means of displacer 8.
  • an amount of washing buffer solution corresponding to the assay is sucked through the distributor 1 and pumped through the outlet hollow body 10.
  • the process can be repeated one or more times as necessary.
  • An optional optimization of the washing process can be achieved by pumping the corresponding wash solution volume into the outlet hollow body 10, removing the magnetic particle fixation by removing the magnetic field supplied outside the hollow body 10 and thus achieving a release of the magnetic particles into the wash solution.
  • a subsequent movement of the newly suspended particles by gently pumping back and forth the resulting suspension by means of displacer and then immobilizing the magnetic Particulate particles by applying a magnetic field is part of a very effective washing process.
  • the excess wash buffer is removed from the outlet hollow body 10 in the next step by drawing in a quantity of gas predetermined by the respective design and subsequent ejection.
  • a necessary for complete detachment of the magnetic particles from the inner boundary of the hollow body 10 amount of substrate is sucked through the manifold 1.
  • the substrate is pumped into the hollow body 10 and brought into contact with the fixed magnetic particles and determined by a detector 12 (PMT, diode, CCD, or the like), the re ⁇ present for the analyte electromagnetic radiation.
  • a detector 12 PMT, diode, CCD, or the like
  • a further optimization of the quantification can be achieved by removing the magnet into the hollow body 10 after the supply of the substrate solution, whereby the previously magnetically fixed particles are released again and by gently pumping back and forth the resulting suspension by means of displacer 8 be mixed with the substrate.
  • the suspension is placed in a detection window or from the hollow body 10 in another vessel.
  • An 8-channel distributor (Hamilton, type N ° 86918) with 100 ⁇ l syringe (Hamilton, type N ° 81022) and Teflon capillaries (Hamilton, type N ° 88907) connected to Luer terminals, microtiter plate white (Nunc, type N ° 437-591), StabilZyme AP (SurModics, Art-N ° SAOI-1000), Streptavidin Magnetic Particles (Aureon Biosystems, Art N ° A010101) and Dynabeads M-280 Streptavidin (Dynal, Art N ° 112.06), Biotin Peroxidase (Sciotec, 5 mg / ml), Peroxidase Substrate (Lumigen, Art-N ° PSA-100), Mediators PhL Luminometer (Aureon, Art-N ° G010001), Neodymoronate Magnet (Magnet Sales, Art N ° NIRD011
  • biotin peroxidase 1: 1,000,000 was diluted with StabilZyme AP using StabilZyme AP as the negative control.
  • the Streptavidin Magnetic Particles were also 1:40 diluted in StabilZyme AP.
  • the liquid was ejected and washed with 25 .mu.l wash solution from a third inlet with applied magnetic field ge. With another 25 ⁇ l washing solution, the particles were washed with the magnet removed. Subsequently, the particles were resuspended with 25 ⁇ l substrate solution, the resulting suspension was ejected into a microtiter plate and measured in the luminometer with 1 second integration time.

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  • Physics & Mathematics (AREA)
  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Immunology (AREA)
  • Pathology (AREA)
  • Investigating Or Analysing Biological Materials (AREA)

Abstract

L'invention concerne un dispositif permettant de déterminer au moins un analyte dans un échantillon, qui comprend un distributeur (1) comportant au moins une ouverture d'entrée (2 à 6) pour introduire des liquides, des suspensions et/ou des gaz dans le répartiteur (1) et au moins une ouverture de sortie (9) pour éliminer des liquides, des suspensions et/ou des gaz hors du distributeur (1). Il est prévu au niveau d'au moins une ouverture de sortie (9) un corps creux (10) tubulaire relié au distributeur (1), corps creux sur lequel est monté un dispositif magnétique (11) pour immobiliser temporairement des particules magnétiques liant les analytes, introduites à l'intérieur du corps creux (10) ou situées à l'intérieur du corps creux (10), au niveau d'une délimitation intérieure du corps intérieur (10).
PCT/AT2005/000477 2004-11-25 2005-11-25 Dispositif de determination qualitative et quantitative d'analytes WO2006056001A1 (fr)

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AT19902004 2004-11-25
ATA1990/2004 2004-11-25

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WO2006056001A1 true WO2006056001A1 (fr) 2006-06-01

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