JP2003501644A - Sample processing equipment - Google Patents

Abstract

(57) Abstract A method for extracting a biological molecule, such as DNA, from a liquid mixture involves passing the liquid up and down along a solid phase containing an adsorbent for the biological molecule.

Description

Detailed Description of the Invention

The present invention relates to sample processing devices, in particular unpurified starting materials (eg blood,
The present invention relates to a sample processing device capable of purifying biological molecules from tissues, plants, microorganisms, agricultural products, foods). The device can also be used to purify or manipulate biological molecules or compounds from aqueous or non-aqueous samples in a fully automated or manual mode.

Conventional chromatographic columns are not suitable for the direct extraction of biological molecules from crude starting materials containing particulate matter, gums, or cell debris. This is due to the type of solid phase used and the design of the column or cartridge which is prone to clogging and clogging. This also applies to mini-chromatography columns that are processed using vacuum or centrifugation to pass the liquid over a solid support. This clogging problem is exacerbated when trying to extract high molecular weight DNA without breaking it into smaller pieces. Clogs in columns or cartridges can be caused by small frit sizes (thick, hard, porous discs / membranes or plugs) or small size standard solid phase particles (typically less than 200 microns in diameter) and / or fixed solids. This may be due to the close packing of the phase material (depending on size and shape, and compressibility).

[0003]   We have devised a device and method that alleviates these problems.

According to the present invention, there is provided an apparatus for extracting a substance from a liquid mixture containing the substance, the container including a solid phase capable of adsorbing the substance to be extracted, and the solid phase. A reversible suction means adapted to exert a suction force to draw up a liquid mixture onto the solid phase, the suction force being reversible so that the liquid reverts from above the solid phase A device is provided.

The reversible suction means may be in the form of a syringe, in which the solid phase is below the piston in the syringe, the nozzle of the syringe is placed in a liquid mixture and the piston is pulled to suck up the liquid onto the solid phase. It is contained so that when pushed the liquid will revert from above the solid phase.

Alternatively, the container containing the solid phase can be attached to the nozzle of a syringe.

In another embodiment of the invention, the reversible aspiration means comprises a pipette, the solid phase stopper being contained within the pipette tip.

A feature of the present invention is that the novel design allows almost all starting materials to be used without clogging the automatic extractor and in a closed environment, to the technician, the instrument or adjacent sample tubes. It is possible to reduce the risk of pollution.

This design allows the use of existing solid phase extraction methods used in chromatography, as well as the new reagents and materials described below.

The present invention is particularly suitable for the extraction of macromolecules such as nucleic acids (DNA and RNA) that are prone to block or clog existing devices.

Syringe or pump systems with suction or blow-out action can be used with specially modified chromatography cartridges that do not clog. For example, a biological sample (eg, animal or plant tissue, blood, cells, hair, feces, produce, water, food) is homogenized to release nucleic acids and then passed through a solid phase material to allow nucleic acids Alternatively, the nucleated cells are captured. Cell debris or contaminants are passed up and down and discarded, leaving the nucleic acid immobilized on the solid support. Alternatively, the solid phase material can be used to leave DNA or biological molecules in solution by removing unwanted cell debris.

The substance to be extracted may be pumped using a syringe or a peristaltic pump.
The solid phase can be passed up and down. Cartridges are designed to be compatible with a wide range of solid phase extractions, so any type of solid phase can be used.

Furthermore, this device allows the homogenization of the sample by introducing a disruption device in the main cartridge.

This is because tedious samples such as plant extracts, animal tissues, faeces, food or other samples that require maceration release cellular contents such as proteins or nucleic acids. To enable. The target molecule can then be captured on the solid phase, thus fully homogenizing and purifying steps.

A concentration step is often used to remove the initial debris, which promotes rapid aggregation and settling of debris or contaminants without the need for cellulose, diatomaceous earth, silica gel, dextran, PEG or centrifugation. Can be performed using an aggregating agent such as any of the substances listed below.

The device of the invention can be designed to operate single or multiple rows of standard syringes (disposable or non-disposable) of various sizes (eg up to 100 ml). A single disposable unit with 4, 8, 12, 24 or 96 channels can also be used with the pump system. That is, it is possible to collect a large amount or a small amount in an array of 8 × 12 tubes.

A feature of the device of the present invention is that the final purified product can be provided in a microtiter plate format of 8 × 12 tubes or as a single tube regardless of the starting volume of the sample. Thus, for the extraction of DNA, a large number of blood samples (eg 5 ml) can be processed at the same time and then concentrated in a small 1 ml tube.

By using aspiration and squirt rather than reagent or sample flow, the disposable cartridge holds the sample completely and at the interface with the device (eg, tubes, valves, and disposable cartridge mouths). Contamination is prevented. This also allows disposable items to be automatically discarded,
Minimize the risk to the technician.

Alternatively, a smaller syringe or pump device can be used to process an entire microtiter plate containing a small sample (eg, 1 ml cheek cell scrape).

A syringe or multi-channel disposable cartridge system
It can be operated in the X, Y and Z directions to accommodate any changes in pitch needed to handle different sample volumes. The purified analyte is UV /
Automatically transferred to a visible spectrophotometer or fluorometer to estimate analyte concentration and purity.

Most conventional UV spectrophotometers require the analysis of relatively large samples in silica-quartz cuvettes. Unfortunately, in biological samples,
The amount of analyte is often low or low. This results in sacrificing the entire sample or diluting it in large amounts, which makes detection very difficult. By providing a disposable probe that is immersed in the sample, the total amount of the solution can be measured without waste.

The design of the device allows the incorporation of electrodes or metal meshes or conductive plastic meshes, which can be positively charged to bind nucleic acids in the crude extract. The purified charge can then be released by turning off or reversing the positive charge. The pumping action of the device allows for rapid mixing and increased target molecule contact. In one mode, the mesh, beads, or chips can be incorporated as a plastic disposable, which can have a positive charge by applying a potential difference or induction. Nucleic acid binds as the sample passes through the mesh or membrane, reversing the charge or switching off, releasing the nucleic acid.

By using very large porous or non-porous solid phase beads, clogging can be avoided and high flow rates can be maintained. Porous diameters greater than 150 microns with very large pores (eg 1-20 microns) made from polypropylene, polyethylene or any polymer with a natural affinity for a particular biological molecule Plastic beads of. Other materials: cellulose, agarose, glass, silica, or any suitable material that can be derivatized to extract the target analyte can also be used.

The solid phase may be derivatized with an imidazole group, an amine, a carboxy, or any group that has an affinity for nucleic acids or target molecules / compounds. The beads can be used with a frit or membrane, or with a single-hole mesh or a multi-hole mesh depending on the flow rate required.

The beads may be composed of substances that have an intrinsic affinity for biological molecules, such as polyvinylpyridine, that are positively charged at pH 4 to bind to DNA and elute it at pH 8. Any polymeric compound can be converted into beads or particles or surfaces for conjugation, including pyrazoles, pyrroles, pyrrolidines, indoles, pyrimidines,
Nucleobase, imidazole, imine, amine, lysine, or pKa in the range 3-12
May contain any group having Preferably, when processing biological samples, pKas 5-8 are used to maintain physiological conditions, whereby the pH can be manipulated to turn positive charges on or off.

The beads may also be converted at pH 3 for chelation of biological molecules such as ferric ion-bound iminodiacetic acid beads to bind nucleic acids. Increasing pH removes purified nucleic acids. Calcium, magnesium or ammonium ions can also be used to chelate biological molecules.

The beads can be immobilized using a frit or membrane as long as the pore size is large enough to allow the unpurified material to pass easily. In most cases, the frit or film requires relatively large holes (pinholes) not found in conventional materials. Alternatively, the beads may be held in place by narrowing the entrance and exit of the cartridge, eliminating the need for frits, or by a mesh built into the cartridge design. Good.

Large beads can be made to move within the column by providing space in the cartridge. This aids mixing and reduces clogging. Alternatively, the beads are added as a suspension in the crude mixture and then, upon binding of the target molecule, are captured in the cartridge, thus effecting the separation.

Large diameter beads are also useful for preparing magnetic solid phases, which do not tend to aggregate during the isolation of macromolecules (eg nucleic acids). Most non-porous magnetic (paramagnetic) beads are less than 50 microns, but are difficult to handle when extracting genomic or microbial DNA. Large porous magnetic beads allow macromolecules such as DNA to bind to the outer surface of the beads, trapping contaminants inside and releasing these contaminants only when the DNA elutes and the final preparation. Pollute.

Porous membranes or frits can be modified by adding more holes, used by stacking at intervals, or biologically separated from the crude extract (eg blood). Used to bind molecules. Existing membranes or frits will occlude immediately upon encountering the crude extract or high molecular weight DNA. The present invention describes the use of a unique pore size modification that allows the processing of large or small amounts of crude material that is also easy to automate.

Small or large devices can be assembled based on the same design for the extraction of DNA from blood. The smaller device relies on a standard plastic pipette tip, which incorporates a single porous stopper, plug, or frit with a pore size that prevents blockage. It can be used with conventional pipettes or syringes in manual mode or with a fully automated pipetting mechanism. Alternatively, they can be incorporated into deep well plates, microtiter plates, or PCR tubes for use with centrifugation or vacuum manifolds. The frit may also be incorporated directly into a 1 ml to 60 ml syringe instead of an additional cartridge.

The stopper or frit may be derivatized to bind DNA or any biological molecule. For larger scale extractions, additional plugs or frits can be added in stacks separated by small air barriers to avoid blockage and maintain an exposed surface for binding the target compound.

The material of the membrane or frit has a main pore size of 1 to 200 microns or preferably 2
0 micron porous polyethylene, or any porous plastic with pores large enough to allow passage of unpurified material (e.g., 20 microns or more),
It may be porous glass, cellulose. The material may have a small pore size to allow binding of target molecules, but it should also have larger pores (eg 0.1 mm or more) to avoid clogging. The larger hole may be at the end of the cut, in the middle, or part thereof.

Various frits can be incorporated into a single cartridge to provide continuous or discrete separations, such as one frit separating nuclei and another derivatized with silica or imidazole groups. DN in one, in cartridge or in another device
Purify A. Alternatively, an initial capture of DNA, RNA, or other biological molecule is made, then the analytes are rinsed, placed in a syringe and precipitated with a compound such as PEG, alcohol, ammonium sulphate or sodium sulphate. The precipitated compound may then be recaptured and the soluble contaminants passed through for disposal.

A syringe-based system can be used to attach a filter membrane or stopper to the tip of a cartridge or pipette tip to filter the crude sample. When the liquid is aspirated through the stopper, debris remains. Flush debris from the stopper and then dispense the clarification solution into a new tube.

This method can be facilitated using filter aids such as silica gel, titanium oxide, fibrous cellulose.

After homogenization, filter aids can be added to agglomerate debris at the bottom of the tube and compressed to leave the target molecule in the supernatant for processing or purification.

The filter aid may be soluble, temperature dependent soluble or insoluble.

When using smaller solid phase beads (eg, less than 100 micron glass) for conventional extraction, bypass channels can be introduced that pass larger particles or debris up and down without clogging the cartridge. . The bypass channel is made as a small tube, which bypasses the solid phase or a porous material with larger pores (eg, 20 microns or more) surrounding the solid phase.

Further purification can be carried out with a separate cartridge inline, which allows the use of salting out methods, in order to obtain very pure DNA. This salting out method can be used with alcohol precipitation and capture of insoluble nucleic acids.

When using smaller solid phase beads (eg, <100 micron glass) for conventional extraction, the solid phase can move inside, thus avoiding clogging. The introduction of ridges, spirals, or obstacles into the cartridge helps prevent the solid phase from clumping and moving, maintaining good mixing and separation of the solid phase. If the solid phase particles are large enough (eg, 200 micron glass) to have sufficient density, a mini-fluidized bed can be created.

Clogging of the cartridge can also be avoided by adding a mobile phase in advance to capture the target molecule. Non-dense solid phase or paramagnetic beads are added as a suspension and the contaminants washed away to leave the immobilized target compound ready for further purification or analysis on a cartridge. Any type of solid phase that has a selectivity for substances that settle rapidly or aggregate with filter aids to avoid centrifugation, or substances that are paramagnetic can be added.

The device and disposable cartridge or chip system have a variety of applications, including molecular biology, such as affinity purification of antibodies, enzymes and other proteins of cells, purging of mixtures to remove unwanted compounds, combinatorial. Chemistry, ion exchange purification, hydrophobic chromatography, immobilized antibodies, enzyme assays using nucleic acids or antigens, enzyme catalysts on solid supports, food screening for pathogens, genomic DNA, toxins, allergens, pathogenic organisms For clinical sample processing, admixture of adjuvants to immunize and create stable emulsions, high volume pipetting, lipoprotein removal for cholesterol assay, detection or concentration of pathogens in milk, food or water There is.

[0044]   The apparatus of the present invention is illustrated in the accompanying drawings below.

[0045]   FIG. 1 illustrates a device having a solid phase in a syringe.

[0046]   FIG. 2 shows the use of the cartridge.

[0047]   FIG. 3 shows a pipette.

[0048]   4, 5, and 6 show another cartridge.

[0049]   7, 8 and 9 show various disk mechanisms.

Referring to FIG. 1, a syringe (1) having a movable piston (2) has an adsorptive solid phase (3) retained therein. In use, the nozzle (4) is placed in the liquid from which the substance is separated, and when the piston is pulled, the liquid is sucked up through (3). When the piston is pushed, the liquid is pushed back from above (3) and this process can be repeated if necessary for better adsorption of the substance from the liquid.

Referring to FIG. 2, a syringe (5) with a piston (6) has a nozzle (7) placed in a cartridge (8) containing a solid adsorbent, the cartridge (8) being It has an inlet (9) contained in the liquid to be separated. When the piston (6) is pulled, the liquid is sucked up through the cartridge, the substance is adsorbed, and when the piston is pushed, the liquid is pushed back over the adsorbent in the cartridge so that the substance is better adsorbed from the liquid. .

Referring to FIG. 4, the adsorbent material (10) may be in the form of frits or beads and may fill the cartridge.

Referring to FIG. 5, a bypass channel (11) is provided on the outer periphery of the solid adsorbent so that larger particles can pass up and down without being clogged.

Referring to FIG. 6, there are discs (12) located in the cartridge, each disc consists of an adsorption membrane, the disc can have large holes as shown in FIG. 7, and as shown in FIG. It may have a cutting portion to prevent blockage. The discs can be stacked on top of each other and can have raised edges (14) as shown in FIG. 9, through which the discs only contact.

With respect to FIG. 3, the pipette (15) has an aerosol plug (16) to prevent contamination and contains a plug (16) of adsorbent material such as a porous plastic material as shown. The tip of the pipette (17) is submerged in the liquid, the liquid is sucked up on the stopper (16), and the liquid is pushed back out of the stopper (16) by blowing out the pipette, resulting in better substance from the liquid. Is adsorbed on.

[0056]   The adsorbed material can be removed from the solid phase by conventional elution methods.

The invention is illustrated in the following examples, in which isolated or eluted products were identified using conventional experimental analytical methods.

Example 1 Using the apparatus of FIG. 2, polystyrene porous carboxylated beads (200-5
00 micron or 16-50 mesh size) was placed in a chromatography cartridge and retained by a plastic mesh with a pore size of approximately 100 microns. 10m whole blood
It was diluted 10 times with M ammonium bicarbonate, 10 mM ammonium carbonate and 0.1% Tween 20 (pH 9), and sucked up and down along the cartridge with a syringe, and then reconstituted through the cartridge. The dilution buffer can be any hypotonic solution that causes lysis of the red blood cell fraction but maintains the integrity of the nucleus, white blood cells or chromatin.
Nuclei were immobilized on beads and lysed blood was discarded. Direct elution of nuclear DNA was performed using warm water. The eluate from the first cartridge was further processed using another cartridge containing a solid phase with polyimidazole groups to obtain higher purity DNA. The solution was made isotonic with saline to collect the white blood cell fraction,
Cells were similarly captured.

Example 2 Using the apparatus of FIG. 3, polystyrene porous carboxylated beads (200-5
00 micron or 16-50 mesh size) was placed in a 1 ml pipette tip. Whole blood was diluted 10-fold with 10 mM ammonium bicarbonate and 0.01% Tween 20 (pH 9) and sucked or sucked along the tip of a pipette. Nuclei were immobilized on beads and lysed blood was discarded. Direct elution of nuclear DNA was performed using alkaline detergent solution and in boiling water.

Example 3 Using the apparatus of Figure 2, agarose was treated with carbonyldiimidazole in anhydrous organic solvent and then left in water at pH 3 to maintain the imidazole group. Derivatized agarose was placed in a cartridge and the supernatant from the plasmid alkaline lysis preparation was aspirated or aspirated to immobilize the plasmid DNA on the beads at pH 5. After washing, the plasmid DNA was eluted with 10 mM Tris-hydrochloric acid (pH 9). The above was repeated with various sizes of carboxylated polystyrene and dextran and eluted as above to obtain DNA.

Example 4 Extraction of Nuclei or DNA from Whole Blood Using the device of FIG. 2 with the packing of FIG. 6, total volume was increased with 5 volumes of 10 mM ammonium bicarbonate containing 0.1% Tween 20 (pH 9). The blood was lysed. Dissolve the lysed blood in a plastic cartridge fitted with a 2 ml syringe and a plunger and have a diameter of 1 mm.
Through several 20 micron porous polyethylene frits modified with larger pores. The frits were placed 3 mm apart to allow the liquid to flow freely. The nuclear or leukocyte fraction bound to the frit and all contaminating proteins and lipids were flushed out and discarded in a single pass or several strokes of the plunger. next,
Wash the frit and core with deionized water or chaotropic agent or alcohol or surfactant (eg SDS or Tween20, or combinations thereof) or lactic acid and salicylic acid or salts thereof, or polyphosphates or perchlorates. Residual proteins were removed and washed with warm water or an alkaline solution of detergent or further purified in cartridge using chaotropic agents or proteases.

Example 5 Purification of buccal cell DNA Using the device of Figure 3, a porous polyethylene plug was derivatized with an imidazole group and inserted into the tip of a standard 1 ml pipette tip. An additional underivatized stopper was inserted on top to act as an aerosol and liquid barrier to prevent contamination of the pipette. A buccal scrape was collected and mixed with 0.2 M guanidine isothiocyanate, 3% Tween 20, proteinase K and 50 mM MES (pH 5).
Mix for 15 minutes at 0 ° C. The mixture was then siphoned or siphoned along the tip to bind the DNA to the derivatized stopper. The stopper was washed with 1 mM MES (pH 5), and then the DNA was eluted with 10 mM Tris-hydrochloric acid (pH 9). The same protocol was repeated in the presence or absence of salt and buffer using 0.01% -10% SDS. 0.05-5
Bacterial cells can also be rapidly degraded using salicylic acid, lactic acid, or MgCl ₂ at concentrations of M. Combinations of the above salts and reagents can also be used.

Example 6 1 g of carboxylated polystyrene beads of about 60 micron diameter or 200-400 mesh were suspended in a hypotonic solution of 10 mM ammonium bicarbonate with 0.1% Tween 20 (pH 9). A 5-fold excess of this suspension was added to 5 ml of blood sample and mixed once. The beads captured the nuclei and settled. After washing with water several times, the DNA was eluted with warm water. D
To concentrate NA, the device of Figure 2 with the packing of Figure 6 was used to capture the DNA on a porous disk in a cartridge, then eluted in a small volume and analyzed by PCR or restriction digestion. did.

Example 7 Collection and Purification of Human IgG from Serum The agarose gel bound to protein A was placed in the cartridge of FIG. 2 and washed with phosphate buffered saline. A solution containing human IgG in serum was added to all I
It was wicked up and down along the solid phase until gG bound. After washing the solid phase with PBS, IgG was eluted with 0.1 M glycine and 0.15 M NaCl (pH 2.8) and immediately neutralized with Tris-hydrochloric acid.

EXAMPLE 8 Purification and isolation of specific leukocyte types from whole blood 175 micron non-porous glass particles were bound to CD4 monoclonal antibody and the solid phase was as in Example 4 with a 80 micron frit. I put it in a cartridge. A diluted solution of Buffy Coat was drawn up and down along the glass beads to immobilize the T cell subpopulation. This could be liberated.

Example 9 Purification of Recombinant Protein The iminodiacetic acid-agarose gel containing nickel ion groups was loaded into the cartridge of Example 4. Bacterial lysates containing recombinant proteins with a 6-histidine tail were siphoned and siphoned along the cartridge to attach the proteins to coordinated nickel. Protein release was performed by elution with 0.5 M imidazole (pH 6).

Example 10 Extraction of HIV RNA from Serum The cartridge of FIG. 2 was filled with 60 micron of silica and diluted 5 times with 6 M guanidine isothiocyanate, 0.1% Tween 20, 20 mM EDTA, 100 mM Tris-hydrochloric acid (pH 6). The sampled serum was drawn up and down along the solid phase. After washing the solid phase with isopropanol and drying, RNA was eluted using warm water at 60 ° C.

Example 11 Purification of PCR Reaction The cartridge of FIG. 2 was filled with 60 micron silica and diluted 5 times with 6M guanidine isothiocyanate, 0.1% Tween 20, 20 mM EDTA, 100 mM Tris-hydrochloric acid (pH 6). A sample of the PCR reaction was sucked up and down along the solid phase. After washing the solid phase with isopropanol and drying, the DNA was eluted using water.

Example 12 Extraction of RNA from Liver Fresh liver was homogenized with a mixture of 50% phenol containing 6M guanidine isothiocyanate, 10 mM DTT, 0.1 M sodium acetate (pH 4). Chloroform was added to separate the phases and the top layer containing RNA was wicked up and down along the cartridge of Figure 2 containing 60 micron silica. The silica was washed with alcohol, air dried, and RNA was extracted with warm water and ready for processing.

Example 13 An mRNA isolation cartridge was filled with oligo dT 30 5'NH 2 conjugated COOH polystyrene beads. Excess 1% SDS of leukocyte sample previously prepared in cartridge,
It was treated with 0.5 M LiCl, 10 mM DTT, 10 mM Tris-hydrochloric acid (pH 7.5), and sucked and sucked several times along the affinity resin to cleave DNA and bind mRNA.
The resin was then washed with 0.1M LiCl and air dried. Elution of mRNA was performed with warm water. The above experiment was repeated using a carboxylated plastic porous frit as in Example 4 conjugated to oligo dT30 and used to bind less than 5 μg of mRNA.

Example 14 Streptavidin immobilized on a solid phase Streptavidin in 0.1M sodium phosphate buffer was mixed with 0.01% glutaraldehyde (pH 7) and mixed on a porous frit as in Example 4. To
Streptavidin was immobilized. The biotinylated primer used to generate the PCR product was isolated on immobilized streptavidin. Then heat or 0.1
The PCR product was made single-stranded using M NaOH and used for sequencing or probe analysis.

Example 15 Use of Electrodes, Electrostatic Charge, Induction, Electrophoresis to Isolate DNA or RNA Whole blood was treated with 10 mM ammonium carbonate / bicarbonate, 50 mM Tris-HCl (with 1% Tween20), 100 lg. Diluted 10 times with / ml proteinase K (pH 9). The electrode was surrounded by a dialysis tube containing the same buffer and immersed in the solution. A 12 volt direct current was connected from the battery, and after incubation for 1 hour, nuclei or DNA was captured outside the dialysis tube with a positive electrode. The DNA could be extracted by elution with water.

[Brief description of drawings]

[Figure 1]   It is a figure which illustrates the apparatus which has a solid phase in a syringe.

[Fig. 2]   It is a figure which shows the use of a cartridge.

[Figure 3]   It is a figure which shows a pipette.

[Figure 4]   It is a figure which shows another cartridge.

[Figure 5]   It is a figure which shows another cartridge.

[Figure 6]   It is a figure which shows another cartridge.

[Figure 7]   FIG. 7 is a diagram showing various disk mechanisms.

[Figure 8]   FIG. 7 is a diagram showing various disk mechanisms.

[Figure 9]   FIG. 7 is a diagram showing various disk mechanisms.

[Procedure for Amendment] Submission for translation of Article 34 Amendment of Patent Cooperation Treaty

[Submission date] June 26, 2001 (2001.6.26)

[Procedure Amendment 1]

[Document name to be amended] Statement

[Name of item to be amended] Claims

[Correction method] Change

[Contents of correction]

[Claims]

─────────────────────────────────────────────────── ─── Continued front page    (81) Designated countries EP (AT, BE, CH, CY, DE, DK, ES, FI, FR, GB, GR, IE, I T, LU, MC, NL, PT, SE), OA (BF, BJ , CF, CG, CI, CM, GA, GN, GW, ML, MR, NE, SN, TD, TG), AP (GH, GM, K E, LS, MW, MZ, SD, SL, SZ, TZ, UG , ZW), EA (AM, AZ, BY, KG, KZ, MD, RU, TJ, TM), AE, AL, AM, AT, AU, AZ, BA, BB, BG, BR, BY, CA, CH, C N, CU, CZ, DE, DK, EE, ES, FI, GB , GD, GE, GH, GM, HR, HU, ID, IL, IN, IS, JP, KE, KG, KP, KR, KZ, L C, LK, LR, LS, LT, LU, LV, MD, MG , MK, MN, MW, MX, NO, NZ, PL, PT, RO, RU, SD, SE, SG, SI, SK, SL, T J, TM, TR, TT, UA, UG, US, UZ, VN , YU, ZA, ZW

Claims (12)

[Claims] 1. An apparatus for extracting a substance from a liquid mixture containing the substance, the container containing a solid phase capable of adsorbing the substance to be extracted, and a suction force acting on the solid phase. A reversible suction means adapted to suck up the liquid mixture onto the solid phase, the suction force being capable of reversing the liquid back over the solid phase. . 2. The apparatus of claim 1, wherein the solid phase contains a syringe, wherein the solid phase is below the piston in the syringe, and the nozzle of the syringe is placed in the liquid mixture and the piston is withdrawn. A device that is sucked up onto and contains a liquid which, when pressed by a piston, returns from the top of the solid phase. 3. The device according to claim 1, wherein the container containing the solid phase is a cartridge attached to a nozzle of a syringe. 4. The device according to claim 1, wherein the reversible means comprises a pipette, the solid phase stopper being contained within the pipette. 5. A device according to any one of claims 1 to 4, wherein the solid phase contains beads adapted to adsorb the substance to be extracted. 6. A device according to any one of claims 1 to 4, wherein the solid phase comprises a plurality of spaced discs. 7. A method for extracting a substance from a liquid mixture, the liquid being drawn up onto a solid phase capable of exerting suction on the solid phase to adsorb the substance to be extracted. A method comprising reversing the suction force to cause the liquid to revert from above the solid phase. 8. The method according to claim 7, wherein the solid phase is contained below the piston in the syringe, the nozzle of the syringe is placed in the liquid, and the piston pulls the liquid onto the solid phase. A method in which the liquid is sucked up, and then the piston is pushed, causing the liquid to flow back from above the solid phase. 9. The method of claim 7, wherein the nozzle of the syringe is placed in the liquid mixture, pulling the piston draws the liquid onto the solid phase, and then pushing the piston solidifies the liquid. The way to go back from the top of the phase. 10. The method according to claim 7, wherein the reversible means comprises a pipette, wherein the solid phase stopper is contained within the pipette. 11. The method according to claim 7, wherein the solid phase contains beads adapted to adsorb the substance to be extracted. 12. The solid phase comprises a plurality of spaced disks.
The method according to any one of claims 7 to 10.