CN212894763U - Device and instrument for nucleic acid extraction and detection - Google Patents
Device and instrument for nucleic acid extraction and detection Download PDFInfo
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- CN212894763U CN212894763U CN201922464802.9U CN201922464802U CN212894763U CN 212894763 U CN212894763 U CN 212894763U CN 201922464802 U CN201922464802 U CN 201922464802U CN 212894763 U CN212894763 U CN 212894763U
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- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6806—Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6844—Nucleic acid amplification reactions
- C12Q1/6851—Quantitative amplification
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Abstract
The utility model provides a nucleic acid extraction and detection device, which comprises one or more sample receiving modules; one or more lysis modules; one or more nucleic acid extraction modules; one or more nucleic acid amplification modules; and a detection module. The utility model also provides a molecular in-vitro diagnostic instrument comprising the nucleic acid extraction and detection device and an automatic control system.
Description
Technical Field
The utility model relates to an external diagnostic device field. In particular, the present invention provides an automated nucleic acid extraction and amplification instrument, and methods of using the instrument for detection and diagnosis.
Background
The method has important effects in extracting bioactive substances, such as cells, and bioactive substances such as proteins and nucleic acids, in various fields such as modern clinical disease diagnosis, transfusion safety, forensic identification, environmental microorganism detection, food safety detection, molecular biology research and the like. Among them, a bioactive substance compound in which nucleic acid is polymerized from many nucleotides is one of the most basic substances of life. Nucleic acids are widely present in all animals, plant cells, microorganisms, and organisms. Nucleic acids can be classified into ribonucleic acids (RNAs) and deoxyribonucleic acids (DNAs) according to their chemical composition. DNA is the primary material basis for the storage, replication, and transmission of genetic information, and RNA plays an important role in the protein synthesis process. With the development of biotechnology, methods and instruments for extracting and detecting nucleic acids from samples are required with the application of PCR technology in various fields, including medical disease detection, agricultural transgene detection, and other applications.
However, at present, regardless of the method, a high-throughput fully-automatic instrument has become a bottleneck for the application of PCR technology in the field of diagnostic analysis. There are several reasons for this. First, many diagnostic analyses are performed with highly specialized equipment only, which is expensive and can only be operated by trained clinicians. Often, in certain areas, there is only one such device. This means that most hospitals need to send samples to these locations for analysis, thus incurring transportation costs and transportation delays, and possibly even leading to sample loss or incorrect operation. Secondly, the equipment in question does not generally run as often as needed, but in batches after a certain number of accumulations, and therefore there is a delay in the processing of many samples, which has a lot of uncertain consequences on the accuracy of the final result.
Therefore, what is needed are nucleic acid extraction and analysis instruments and methods that are as automated as possible, while being suitable for ready-to-run use, and that are better able to avoid contamination. For example, once extracted from a patient, a biological sample must be placed in a form suitable for PCR processing to amplify a vector of interest, such as a nucleotide. Once amplified, the nucleotide of interest from the sample needs to be clearly determined. Also, the instrument should have a high throughput, especially for one sample to perform a number of different analyses; but also in a manner that is routinely performed at the point-of-care, without the need to send the sample to specialized equipment for testing.
SUMMERY OF THE UTILITY MODEL
The device and instrument provided by the utility model have high flux to can carry out nimble distribution to the flux according to the demand, except can detecting a plurality of samples simultaneously, also can carry out the test analysis of multiple difference to a sample.
Specifically, the utility model provides a nucleic acid extraction and detection device, the device includes:
one or more sample receiving modules, each sample receiving module having a compartment adapted to receive a sample or sample container;
one or more lysis modules, each lysis module having a compartment adapted to receive a lysis reagent or a lysis kit;
one or more nucleic acid extraction modules, each nucleic acid extraction module having a compartment adapted to receive a nucleic acid extraction reagent or nucleic acid extraction kit;
one or more nucleic acid amplification modules, each nucleic acid amplification module having a plurality of compartments adapted to receive nucleic acid amplification reagents or nucleic acid amplification kits; and
and a detection module.
In one aspect of the present invention, the sample receiving module in the device has a sample rack capable of accommodating sample tubes, and is disposed at the front end of the device close to the operator.
In one aspect of the present invention, the sample is lysed in a lysis module of the device by using a lysing agent, an enzyme, ultrasound, or physical grinding.
In one of its aspects, the lysis module in the device also has an element for lysing the sample.
In one aspect of the present invention, the component of the cracking sample in the device is an oscillator, which is composed of an eccentric shaft, an eccentric seat and a motor, wherein the eccentric shaft rotates around the axis of the motor, so that the eccentric seat oscillates.
In one aspect of the invention, the lysis module in the device is adapted to receive abrasive particles or a lysis kit comprising abrasive particles to lyse the sample in a physical abrasion manner.
In one aspect of the present invention, the lysis kit has a lysis tube that can hold abrasive particles. In one aspect of the invention, the cracking tube has a cover and the inner wall has two or more axial strip-like projections.
In one aspect of the invention, the lysis module in the device is adapted to receive a lysis agent or a lysis kit comprising a lysis agent to chemically or biologically lyse the sample.
In one aspect of the present invention, the nucleic acid extraction module in the device has one or more nucleic acid extraction units. In yet another aspect of the present invention, the nucleic acid extraction unit is comprised of a binding unit, a washing unit, and an elution unit.
In one aspect of the present invention, the nucleic acid extraction module in the device has a magnet device that can apply a magnetic field, control magnetic material and/or bind the magnetic material of nucleic acid.
In one aspect of the present invention, the device has a dispensing tube that can be inserted into the combination unit, the cleaning unit and the elution unit, and the nucleic acid in the container is extracted by the magnetic bead method, wherein the nucleic acid sample is combined with the magnetic bead in the solution of the container, the tip of the dispensing tube can be inserted into the bottom of the container, the magnet is close to the dispensing tube, the magnetic bead and the combined sample are adsorbed to the side wall of the dispensing tube by the magnet, the dispensing tube moves to another container after the liquid is discharged, the magnet leaves the dispensing tube, the magnetic field effect disappears, and the adsorbed magnetic bead and the sample are released.
In one aspect of the present invention, the nucleic acid amplification module in the device is configured to be suitable for performing isothermal amplification or PCR, preferably, it is suitable for fluorescent quantitative PCR of DNA and RNA.
In one aspect of the present invention, in the nucleic acid amplification module in the device, the nucleic acid amplification module has a plurality of nucleic acid amplification units. Each nucleic acid amplification unit generally corresponds to a nucleic acid extraction sample obtained by each nucleic acid extraction unit in the nucleic acid extraction module. The nucleic acid amplification unit corresponding to the nucleic acid extraction sample obtained by each nucleic acid extraction unit in the nucleic acid extraction module comprises compartments for performing two or more amplification reactions. In one aspect of the present invention, each of the nucleic acid amplification units may perform two or more identical or different amplification reactions on the same nucleic acid extraction sample.
In one aspect of the present invention, the nucleic acid amplification module in the device further comprises a temperature regulator that can independently heat or cool each of the plurality of compartments, such that each compartment can independently perform a different or the same amplification reaction.
In one aspect of the present invention, the nucleic acid amplification module in the device further comprises a flip mechanism to close the lid of the amplification tube. In one aspect of the present invention, the flip mechanism has a cover pressing component and a moving component, wherein the cover pressing component includes a cover pressing plate and a stepping motor, and the stepping motor controls the cover pressing plate to move up and down; the motion assembly can move back and forth in the horizontal direction, and the amplification compartment of the nucleic acid amplification module is connected with the motion assembly; the cover turning mechanism is arranged in a way that when an amplification tube in an amplification compartment or a nucleic acid amplification kit moves to be close to the cover pressing plate along with the movement component, the stepping motor drives the cover pressing plate to move upwards, the cover of the amplification tube is slowly lifted, the tube cover is turned over, when the cover of the amplification tube is lifted to a proper angle, the cover pressing plate does not move upwards any more, the movement component continues to move towards the cover pressing plate, and then the stepping motor drives the cover pressing plate to move downwards, so that the tube cover is pressed.
In one aspect of the invention, the detection module in the device detects the identifiable label carried by the nucleic acid, including but not limited to fluorescence or other forms of luminescence (e.g., chemiluminescence, bioluminescence, radioluminescence, electroluminescence, electrochemiluminescence, mechanoluminescence, crystallography, thermoluminescence, sonoluminescence, phosphorescence, photoluminescence, and the like), enzymatic reactions, radioactivity, and the like.
In one aspect of the invention, the detection module in the device comprises a fluorescence analyzer.
In one aspect of the present invention, the device further comprises a liquid dispensing module to transfer or dispense a sample, reagent or other liquid in the device between two or more locations.
In one aspect of the present invention, a liquid dispensing module in the device comprises: one or more sensors; one or more dispensing heads; and a displacement control mechanism providing three-dimensional translational motion freedom for the dispensing head.
In one aspect of the present invention, the device further comprises a reagent and consumable setting area.
In one aspect of the present invention, the reagent and consumable material setting area in the apparatus includes one or more of a pipette tip setting area, a PCR reagent storage and mixing processing area, and a waste material area.
The utility model also provides an automatic molecular in vitro diagnostic apparatus, it includes aforementioned nucleic acid extraction and detection device to and for example automatic control system based on computer.
In one aspect of the present invention, the diagnostic device is used for infection source identification, genetic disease, cancer detection, or genetic variation detection.
In one aspect of the present invention, the diagnostic device is used for the detection of the following pathogens: influenza virus, enterovirus, hepatitis B virus, hepatitis C virus, Ebola virus, Marburg virus, SARS virus, Securia virus, bunyavirus, rhinovirus, respiratory nucleocapsid virus, cholera virus, etc., or bacterial pathogens such as tubercle bacillus, Escherichia coli, Acinetobacter baumannii, Diplococcus pneumoniae, Streptococcus lactis, Sporosarcina urens, Staphylococcus aureus, Bacillus subtilis, Bacillus anthracis, Bacillus subtilis, Bacillus streptococci, Bacillus proteus, Vibrio cholerae, Treponema pallidum, etc.
In one aspect of the present invention, the diagnostic device is used for the detection of the following cancers: gastric cancer, liver cancer, lung cancer, esophageal cancer, cervical cancer, breast cancer, colon cancer, rectal cancer, nasopharyngeal cancer, ovarian cancer, renal cancer, bladder cancer, thyroid cancer, skin cancer, etc.; malignant tumors derived from mesenchymal tissues such as muscle, fat, bone, blood vessel, lymph, etc., for example, rhabdomyosarcoma, leiomyosarcoma, fibrosarcoma, liposarcoma, osteosarcoma, chondrosarcoma, angiosarcoma, lymphosarcoma, etc. Also, for example, leukemia, Hodgkin's disease, Wilms ' tumor (nephroblastoma), melanoma, retinoblastoma, seminoma, granuloma, Kupffer's tumor, Ewing's tumor, malignant vascular endothelial cell tumor, or Paget's disease of the breast.
Drawings
Other features, objects and advantages of the invention will become more apparent upon reading of the detailed description of non-limiting embodiments made with reference to the following drawings:
fig. 1 is a perspective view of an exemplary device of the present invention.
Fig. 2 is a schematic view of a partial plan layout of an exemplary apparatus of the present invention.
Fig. 3 is a block diagram of an exemplary sample holder of the present invention.
Figure 4 is a block diagram of an exemplary lysis module of the apparatus of the present invention.
Figure 5 is an exemplary lysis kit of the present invention.
Fig. 6 is a schematic diagram of the nucleic acid extraction module using the magnetic bead method and the method for magnetic field controlled adsorption pipetting in the module according to the present invention.
FIG. 7 is a diagram showing the structure of an exemplary nucleic acid extraction kit of the present invention.
Fig. 8 is an exemplary flip mechanism of the inventive apparatus.
FIG. 9 is a block diagram of an exemplary nucleic acid amplification kit of the present invention.
Fig. 10 is a schematic diagram of an exemplary fluorescence analyzer configuration and optical path of the inventive device.
Fig. 11 is a schematic diagram of an exemplary liquid dispensing module of the apparatus of the present invention.
Figure 12 shows a cross-sectional view of an exemplary single arm pipette of the inventive apparatus.
Fig. 13 is a schematic diagram of another exemplary liquid dispensing module of the apparatus of the present invention.
Detailed Description
The term nucleic acid as used herein includes DNA (deoxyribonucleic acid) and RNA (ribonucleic acid). It is also to be understood that the terms nucleic acid and polynucleotide may be used interchangeably herein.
It is to be understood that the devices described herein have application to the analysis of any nucleic acid-containing sample for any use, including but not limited to genetic testing for human genes and clinical testing for various infectious diseases. The nucleic acid sample used in the methods described herein can be from any source. In general, the sample may be a biological material that is separate from its natural environment and contains polynucleotides. The sample may consist of purified or isolated polynucleotides, or may comprise a biological sample such as a tissue sample, a biological fluid sample, or a cell sample comprising polynucleotides. Biological fluids include, as non-limiting examples, blood, plasma, sputum, urine, cerebrospinal fluid, lavage fluid samples. The nucleic acid sample may be from a plant, animal, bacterial or viral source. Samples can be obtained from different sources, including but not limited to samples from different individuals, different developmental stages of the same or different individuals, different diseased individuals (e.g., individuals with cancer or suspected of having a genetic disorder), normal individuals, different disease stages of the same or different individuals, individuals treated for different diseases, individuals under different environmental factors, or individuals with a predisposition to a disease, or individuals exposed to an infectious disease agent (e.g., HIV).
The term "unit" as used herein is intended to mean an element or combination of elements which are configured to operate together to perform one or more functions or produce one or more desired results, wherein each element may have separate, distinct, and/or independent functions. It will be understood that each element within a unit need not be directly connected or in direct communication with each other element. Also, the connection or communication of the plurality of elements may be realized with the aid of elements external to the unit, such as a processor.
The term "module" is used herein to denote a unit or combination of units configured to operate together to implement one or more subsystem functions of the inventive apparatus. It will be understood that each element within a module need not be directly connected or in direct communication with each other element. Furthermore, the connection or communication of the units may be implemented with the aid of units or elements external to the module, such as a processor.
The "magnetic material" described in the present specification includes various materials having magnetism, including particles and particulate materials having various particle size distributions. The shape of the material is not limited to a spherical material, and any shape is allowable.
The utility model provides a nucleic acid in the sample is used for separating or obtaining to it carries out the device that detects through polymerase chain reaction.
For a better understanding and explanation of the present invention, the present invention will be described in further detail below with reference to the accompanying drawings.
See fig. 1 and 2. Wherein fig. 1 is a perspective view of an exemplary device of the present invention, and fig. 2 is a schematic view of a partial plan layout of an exemplary device of the present invention.
As shown in fig. 1, the device of the present invention comprises:
one or more sample receiving modules 1 having compartments adapted to receive samples or sample containers;
one or more lysis modules 2 having compartments adapted to receive a lysis kit; in the lysis module of fig. 1, there is also an element for lysing the sample;
one or more nucleic acid extraction modules 3, each nucleic acid extraction module having a compartment suitable for receiving a nucleic acid extraction kit comprising a binding unit, a washing unit, an elution unit, and a magnet device movable relative to each binding unit, washing unit or elution unit;
one or more nucleic acid amplification modules 4, each nucleic acid amplification module having two or more compartments adapted to receive a nucleic acid amplification kit;
a detection module 5; and
a liquid dispensing module 6 to transfer or dispense a sample, reagent or other liquid in the device between two or more locations.
Sample receiving module
In the present invention, the device of the present invention has a sample receiving module 1 having a compartment suitable for receiving a sample or sample container. In one embodiment of the present invention, the compartment adapted to receive a sample container is a sample rack 11. An exemplary sample holder may be seen in fig. 3, which includes a holder 12 and a sample tube well 13 for insertion of sample tubes.
An example of a sample container is a sample tube. The sample tube may be a standard sample tube, such as a standard blood collection tube or a BD sample tube. Sample tubes suitable for the device of the present invention may be any commercially available sample tubes, for example, but not limited to, 12/13X75mm glass blood collection tubes, 12/13X75mm plastic blood collection tubes, 13X100mm blood collection tubes, 16X75mm blood collection tubes, 16X100mm blood collection tubes, 16X100mm BD sample tubes, and the like. The sample tube may also be a non-standard sample tube. The type of sample tube selected depends on the consumables and the structural design of the sample tube rack. In the apparatus and method of the present invention, the minimum sample loading of the sample tube can be as high as 500ul for samples that do not require additional diluent addition. For samples that require the use of diluent (or a small sample volume), the minimum sample loading of the sample tube may be 100ul or less. In the example shown in fig. 3, a row of one sample rack can accommodate 12 sample tubes. The sample frame can be arranged at the front end of the device, close to the operator, of the experimental area, so as to facilitate the operation.
In one example of the present invention, the sample rack or the sample tube is further attached with a barcode. The barcode may be scanned externally (e.g., using a handheld laboratory (clinical laboratory) information system), by an internal scanner, or by an instrument-internal automated scanning device, thereby recording and monitoring the sample detected.
Lysis module
The device of the present invention is used for obtaining the nucleic acid of a sample and for performing a method of detection, the device of the present invention comprises a lysis module 2 for lysing the cells of the sample so that the nucleic acid coated by the cell membrane is released.
As shown in fig. 1 and 2, the lysis module 2 of the device of the present invention has a compartment adapted to receive a lysis reagent or a lysis kit. The utility model discloses the mode schizolysis sample that schizolysis agent, enzyme, supersound or physics ground can be adopted to the schizolysis module of device. Wherein lysis reagents (including various reagents and/or solutions used to lyse cells of a sample) are added to lyse the cells. The lysis reagent may also be provided in the form of a lysis kit (e.g., a lysis strip) containing the lysis reagent.
In one embodiment of the present invention, the lysis module lyses the sample using a lysing agent. The utility model discloses usable cracking agent includes various surfactants such as SDS, Triton X, NP-40 etc. and other chemical reagent such as buffer, protease inhibitor, reductant etc. and its main function is: (1) disrupting the lipid bilayer using detergent, disrupting the cells; (2) dissolving the protein; (3) promoting protein denaturation; (4) inhibiting the activity of protease and nuclease. In this embodiment of the invention, the element for lysing the sample is an element that facilitates lysis of the sample by the lysing agent, including but not limited to an optional heating element, a shock generator that facilitates mixing of the lysing agent with the mixture of the sample, or a repeatedly aspirated dispensing tube, etc. The cracking agent is contained in the cracking kit of the utility model, and can be connected with or disconnected with the cracking tube in the cracking tube or other containers of the cracking kit.
In one embodiment of the present invention, the lysis module lyses the sample using enzymatic lysis. The utility model discloses usable enzyme can be the enzyme of various schizolysis cell walls or cell membrane mesogens, for example labase lyase, lysostaphin, hen egg albumen source lysozyme, humanized lysozyme, achromopeptidase, the streptococcal source of ball spore becomes lysostaphin, staphylococcus aureus source alpha-hemolysin, chitinase, rhizoctonia solani source lyase, arthrobacter luteus source lyase, trichoderma harzianum source lyase, streptococcus pyogenes source hemolysin O, tetanus bacillus source tetanus bacteria hemolysin etc.. In this embodiment of the invention, the element for lysing the sample is an element that facilitates enzymatic lysis of the sample, including but not limited to an optional heating element, a shock generator that facilitates mixing of the enzyme with the mixture of the sample, or a repeatedly aspirated dispensing tube, and the like. The enzyme may be in the lysis kit of the present invention, may be in the lysis tube, or in another container of the lysis kit, and may or may not be connected to the lysis tube.
In an embodiment of the present invention, the sample is cracked by the cracking module in an ultrasonic cracking mode. Ultrasonic lysis is the process of breaking cells by ultrasonic heating. However, this treatment easily causes the fragmentation of DNA, so the sonication time and the gap time should be set to avoid the destruction of the target DNA. In this embodiment of the invention, the element for lysing the sample is, for example, an sonicator, and optionally a heating or cooling element to maintain the temperature of the sample. Optionally, the sample is sonicated after mixing with a diluent or buffer. The diluent or buffer may be in the lysis kit of the present invention, may be in the lysis tube, or in other containers of the lysis kit, and may or may not be connected to the lysis tube.
In an embodiment of the present invention, the sample is lysed by the lysis module using physical grinding. Physical grinding means that fine particles having a certain hardness, i.e., abrasive particles such as glass beads or porcelain beads, are mechanically crushed by being vortexed and oscillated together with a sample. Which uses the rapid motion of abrasive particles to impact sample cells to mechanically disrupt and lyse the sample cells. Compared with other modes, the physical grinding is a powerful cracking method, has higher cracking efficiency, can crack cells more uniformly and comprehensively, and has higher DNA yield.
In the lysis module of the exemplary device of the present invention as shown in fig. 4, there are compartments adapted to receive a sample tube or lysis kit of lysis reagents and elements for lysing the sample by physical abrasion. In this embodiment of the present invention, the element for lysing the sample is a shaker 21. As shown in the left drawing of fig. 4, the oscillator 21 rotates around the axis of the dc motor 23 by using the eccentric shaft 22, so that the eccentric seat 24 generates high-speed vibration. The sample container is placed in the eccentric seat, and the glass beads or the ceramic beads added in the sample move at high speed and vibrate, so that the purpose of breaking cells in the sample is achieved. The exemplary lysis module shown in the right diagram of fig. 4 has 12 oscillators, which can process 12 samples simultaneously, and can drive 12 eccentric seats by 12 independent motors, so that the sample in the sample tube or lysis kit with lysis reagent added, which is placed in the eccentric seats, can rotate and collide with the glass beads or ceramic beads therein at high speed. The motor used in the device of the present invention and its rotation speed can be determined by a person skilled in the art according to the prior art or limited experiments.
In the exemplary devices of the present invention, a lysis kit may be employed in the lysis module. In the above-described embodiment of the above-described mode schizolysis sample that adopts physical grinding of the utility model, the schizolysis kit has the fixed or detachable lysis pipe that holds sample/schizolysis reagent.
The lysis kit further comprises abrasive particles, such as glass or ceramic beads for grinding, having a particle size of about 0.1-1 mm. The abrasive particles may be placed in the lysis tube or in another container of the lysis kit that may or may not be connected to the lysis tube. The lysis kit has an outer membrane on the surface that encloses the abrasive particles or lysis reagent within the lysis kit. At the during operation, the operating personnel arranges the schizolysis kit in after tearing the adventitia on schizolysis kit surface the utility model discloses the corresponding compartment of the schizolysis module of device.
In the exemplary lysis kit 25 shown in fig. 5, it comprises 12 lysis tubes 26, wherein each lysis kit can be used for lysis of one sample, as shown in the left panel of fig. 5. The right drawing of fig. 5 is a cross-sectional view of the lysis tube 26 showing said tube 26 having a closure 28 with at least two symmetrical axial strip-like protrusions 27 on its inner wall.
In an embodiment of the present invention, the sample added to the device of the present invention does not need to additionally add diluent. In this case, the minimum sample size may be 50. mu.l, or 100. mu.l, or up to 500. mu.l.
Nucleic acid extraction module
The utility model discloses a device is used for obtaining the nucleic acid of sample and the method that detects carries out, the utility model discloses a device includes nucleic acid extraction module 3, and it is arranged in with the nucleic acid separation in the sample and obtains. In one embodiment of the present invention, after the sample is lysed, the liquid distribution module transfers the lysed sample from the lysis module, e.g., the aforementioned lysis kit, to the nucleic acid extraction module.
As shown in fig. 1 and 2, the nucleic acid extraction module 3 of the device of the present invention has a compartment adapted to receive a nucleic acid extraction reagent or nucleic acid extraction kit. The utility model discloses nucleic acid is drawed through nucleic acid combination, washing and elution step to the nucleic acid extraction module of device, wherein need add nucleic acid combination, nucleic acid washing and nucleic acid elution reagent (including nucleic acid extraction magnetic bead, washing liquid, eluant etc.). Nucleic acid binding, nucleic acid washing and nucleic acid eluting reagents may also be provided in the form of a nucleic acid binding kit (e.g., a nucleic acid binding strip) containing such reagents.
In the method and apparatus of the present invention, the extraction of nucleic acid is performed by a magnetic bead method. Wherein the binding of the nucleic acids is performed by contacting a magnetic material capable of binding to the nucleic acids and/or a binding solution with the sample having undergone the lysis step, thereby allowing the nucleic acids to bind to the magnetic material. The complex formed after the nucleic acid is combined with the magnetic material can controllably move, stir or precipitate in the container under the action of a magnetic field, so that the purposes of nucleic acid combination, nucleic acid cleaning and nucleic acid elution are achieved.
As shown in the schematic partial plan layout of the exemplary apparatus of fig. 2, in one embodiment of the present invention, the nucleic acid extraction module of the apparatus of the present invention has 12 nucleic acid extraction units 31, each of which is composed of a binding unit 32, a washing unit 33, and an elution unit 34, as shown in fig. 2. Each nucleic acid extraction unit is used for processing a sample to obtain a nucleic acid extraction sample. The binding unit 32 of the nucleic acid extraction module is a well that contains a magnetic material and/or a binding solution that can bind to nucleic acids, or is adapted to receive a corresponding binding tube in a nucleic acid extraction kit, which contains a magnetic material and a binding solution that can bind to nucleic acids. When adopting the nucleic acid extraction kit, put into at the nucleic acid extraction kit the utility model discloses add the combination pipe by liquid distribution module behind the device. The washing unit 33 is a plurality of wells for receiving washing solutions or corresponding washing tubes in the nucleic acid extraction kit. When adopting the nucleic acid extraction kit, put into at the nucleic acid extraction kit the utility model discloses add the scavenge pipe by liquid distribution module behind the device. The elution unit 34 is a well adapted to receive an elution solution from a corresponding elution tube in the nucleic acid extraction kit.
The nucleic acid extraction module includes a magnet for moving a magnetic material and nucleic acids bound to the magnetic material between the binding unit, the washing unit, or the elution unit by a magnetic field.
FIG. 6 is a schematic diagram illustrating a nucleic acid extraction module using a magnetic bead method, and a method for controlling adsorption pipetting by a magnetic field in the module. In the magnetic field controlled adsorption pipetting mechanism and method of the exemplary magnetic bead method shown in fig. 6, after the lysed sample is combined with the magnetic beads in the combining tube 35, the tip of the dispensing tube 39 is inserted into the bottom of the magnetic bead combining tube, the magnet 38 is close to the dispensing tube, and the magnetic beads are adsorbed to the side wall of the dispensing tube by the magnet during the slow pipetting up process. After the sample liquid is completely absorbed, the distribution pipe and the magnet synchronously move upwards, then the liquid is discharged, after the side wall adsorption action is completed, the distribution pipe is moved to the cleaning pipe 36 by the horizontal movement of the mechanical arm, the magnet leaves the distribution pipe, the magnetic field action disappears, the adsorbed magnetic beads and the sample are released, the tip end of the gun head of the distribution pipe is inserted below the liquid level, and the magnetic beads enter the cleaning pipe through up-down repeated adsorption and discharge. Similarly, the magnetic material in the wash tube can be transferred to the elution tube 37 by the same method, and the magnetic material is taken into the eluent in the elution tube. After the elution is finished, the magnetic material which is not combined with the nucleic acid is adsorbed to the side wall of the distribution pipe by applying a magnetic field, so that the magnetic material is separated from the eluent.
Similarly, the magnetic material in a wash tube may be transferred to another wash tube, or to an elution tube, in a similar manner. In one embodiment of the invention, each nucleic acid extraction module comprises a plurality of washing units, e.g.2-5, preferably 3. Correspondingly, each nucleic acid extraction kit comprises a plurality of washing tubes, for example 2-5, preferably 3.
In one embodiment of the invention, the magnet arrangement is arranged to be movable to the side near the dispensing tube. In one embodiment of the invention, a magnet arrangement is provided fixed to the dispensing tube of the liquid dispensing module, for example an electromagnet, which applies a magnetic field to the dispensing tube by periodically switching on and off. In one embodiment of the invention, the magnet means, e.g. a permanent magnet or an electromagnet, is arranged to be movable relative to the dispensing tube, e.g. close to or away from the dispensing tube, thereby periodically applying a magnetic field to the dispensing tube. The magnet device may be moved together with the dispensing tube, e.g. closer to or further away from the binding unit, the washing unit or the elution unit, in order to transfer the magnetic material from one unit to another. In this way, the magnet arrangement periodically applies a magnetic field to the dispensing tube, which magnetic field is applied to accumulate magnetic material in the liquid dispensing module, e.g. dispensing tube, and which magnetic material is removed from the liquid dispensing module, e.g. dispensing tube, to transfer to the other modules.
In one embodiment of the present invention, after the sample is lysed, the liquid distribution module transfers the lysed sample from the lysis kit to the nucleic acid extraction kit located in the nucleic acid extraction module. For example, the liquid dispensing module draws the lysed sample into a dispensing tube, then the displacement unit moves the dispensing head and dispensing tube to the nucleic acid extraction cartridge, and finally discharges the lysed sample into the nucleic acid extraction cartridge, e.g., a binding tube adapted for the binding unit.
According to the present invention, the lysed sample is sucked into the dispensing tube by the liquid dispensing module and then displaced in the dispensing tube to the binding unit in the nucleic acid extraction module, e.g. in the binding tube of the nucleic acid extraction kit located in the binding unit. The lysed sample is bound to a magnetic material in a binding tube. After a period of bonding, the dispensing tube tip is inserted into the bottom of the bonded tube. Then, the magnet device applies a magnetic field to the dispensing tube so that the magnetic material combined with the nucleic acid is attracted to the side wall of the dispensing tube by the magnetic field during the slow pipetting-up process of the dispensing tube. When the liquid is completely absorbed, the distributing pipe and the magnet device synchronously move upwards, and then the liquid without the magnetic material is discharged out of the distributing pipe. After the side wall adsorption action is completed, the liquid distribution module horizontally moves the distribution pipe to the cleaning pipe positioned in the cleaning unit, and then the magnet device removes the magnetic field applied to the distribution pipe, so that the magnetic material is released. When the tip of the distribution pipe is inserted under the liquid level, the liquid distribution unit drives the liquid in the distribution pipe to repeatedly enter and discharge the liquid out of the distribution pipe, so that the magnetic material enters the cleaning liquid in the cleaning pipe.
Finally, the liquid dispensing module moves the dispensing tube with the magnetic material adsorbed thereon away from the elution unit, leaving only the elution solution containing nucleic acids in the elution tube. The distribution pipe with the magnetic material adsorbed thereon may be released from the liquid distribution module to the waste region.
Preferably, the binding unit, the washing unit and the elution unit each include a heating element that heats the reaction reagent (e.g., binding or elution solution) and the magnetic material to facilitate binding or separation of the nucleic acid to the magnetic material.
In the above exemplary apparatus of the present invention, a nucleic acid extraction kit may be used in the nucleic acid extraction module. The nucleic acid extraction kit includes a binding tube, one or more washing tubes, and an elution tube, and contains a magnetic material that can bind to nucleic acids, a washing solution, an elution solution, and the like, respectively. In the exemplary nucleic acid extraction kit 31 shown in fig. 7, it may be a single channel reagent strip. The left figure of FIG. 7 is a front view of the nucleic acid extraction kit, and the right figure of FIG. 7 is a perspective view. The nucleic acid extraction kit of the utility model can be also packaged in multiple ways, for example 12 pieces of packaging, wherein each nucleic acid extraction kit can carry out the nucleic acid extraction of a lysis sample.
The binding tube 35, the washing tube 36 and the elution tube 37 in the nucleic acid extraction kit have sealing films. This membrane can adopt two membrane structures, and the inner membrance wherein is the rubber or the plastic envelope membrane that have the cross appearance, and the adventitia is plastic-aluminum or plastic envelope membrane, prevents that the membrane structure prevents liquid external splash in the reagent transportation, and the piping is moved the liquid through the application of sample of cross department during the liquid-transfering. The kit can be pasted with a bar code which contains information such as reagent expiration date.
Nucleic acid amplification module
The device of the utility model is used for the method of amplifying and detecting the nucleic acid of sample, the utility model discloses a device includes nucleic acid extraction module 4, and it is used for the nucleic acid amplification in the sample. In one embodiment of the present invention, after nucleic acid extraction, the liquid distribution module transfers the sample from the nucleic acid extraction module, e.g., the aforementioned nucleic acid extraction kit, to the nucleic acid amplification module.
In one embodiment of the present invention, the nucleic acid amplification module is a module suitable for performing nucleic acid amplification in any manner, including but not limited to various isothermal amplifications or PCRs, such as qPCR (fluorescent quantitative PCR), RT-PCR, hot start PCR, nested PCR, multiplex PCR, reconstitute condition PCR, dsRNA synthesis, COLD-PCR, digital PCR, and the like. Preferably, the nucleic acid amplification module is configured for fluorescent quantitative PCR of DNA and RNA.
As shown in fig. 1 and 2, the nucleic acid amplification module 4 of the device of the present invention has a plurality of compartments 41 adapted to receive nucleic acid amplification reagents or nucleic acid amplification kits. Each of the compartments may be subjected to a separate amplification reaction. In one embodiment of the present invention, in the nucleic acid amplification module, the nucleic acid amplification unit corresponding to the nucleic acid extraction sample obtained by each nucleic acid extraction unit in the nucleic acid extraction module includes compartments for performing two or more amplification reactions, i.e., each nucleic acid amplification unit can perform two or more identical or different amplification reactions on the same nucleic acid extraction sample. In one embodiment of the present invention, each nucleic acid amplification unit corresponding to the nucleic acid extraction sample obtained from each nucleic acid extraction unit in the nucleic acid extraction module has 2 to 30 compartments, preferably 3 to 10 compartments, and most preferably 4 to 8 compartments.
As shown in the schematic partial plan layout of the exemplary apparatus of fig. 2, in one embodiment of the present invention, the apparatus of the present invention has 12 nucleic acid extraction units 31, each corresponding to one amplification unit 45 (i.e., the exemplary apparatus has 12 amplification units 45), which have 4 amplification compartments 41, each of which can perform an independent amplification reaction. The templates of the amplification reactions carried out by the plurality of amplification compartments in one amplification unit are all from the extracted sample provided by the same nucleic acid extraction unit; the amplification reactions of the several amplification compartments in each amplification unit are independent, e.g.the same or different primer pairs may be used, or different PCR reaction conditions, e.g.temperature, may be used.
In the above exemplary apparatus of the present invention, a nucleic acid amplification kit comprising two or more amplification tubes and a premixed reagent containing all the components required for the nucleic acid amplification reaction may be used in the nucleic acid extraction module. The premixed reagent may be placed in the amplification tube, or may be transferred to the amplification tube through the liquid distribution module in an additional container connected or disconnected to the amplification tube. Preferably, each nucleic acid amplification kit comprises 2-30 amplification tubes, preferably 3-10, most preferably 4-8.
In the exemplary nucleic acid amplification kit 43 shown in FIG. 8, it is in the form of a 4-up tube comprising 4 amplification tubes 431 each in the form of a reaction cup with a lid 432, and the bottom end of the cup body is in a tapered configuration and the upper end is in a cylindrical shape. The effective volume of the amplification tube is generally about 1 to 100. mu.l, preferably about 10 to 50. mu.l, and more preferably about 30. mu.l.
The present inventors have found that the challenge of using PCR as the primary method of diagnosis is the presence of a variety of possible pathogenic microorganisms and low levels of microorganisms in certain samples. Running a large number of PCRs is often impractical and in some cases may not have enough samples to test for all possible pathogens. The present invention is based on the idea of inventively including two or more compartments in each nucleic acid amplification module suitable for receiving nucleic acid amplification kits and using nucleic acid amplification kits comprising two or more amplification tubes, so that multiplex PCR analysis can be performed simultaneously on one sample, thereby detecting multiple targets. This not only reduces the sample size requirements, but also increases system throughput in similar configurations, thereby reducing the cost of testing.
In one embodiment of the present invention, the nucleic acid amplification module further comprises a temperature regulator that can independently heat or cool each of the plurality of compartments 41, such that each compartment can independently complete a different or the same amplification reaction. In one embodiment of the present invention, the nucleic acid amplification module operates at a constant temperature and the temperature regulator maintains the temperature of the various compartments of the device. In another embodiment of the present invention, the temperature regulator cycles heating and cooling each compartment of the device in a predetermined program for a predetermined time. The temperature, duration and number of cycles of heating and cooling can be determined by one skilled in the art based on prior art or limited experimentation, and all such embodiments are included within the scope of the present invention.
In the PCR reaction, the amplification reaction of the sample must be performed in a closed environment. The nucleic acid amplification compartment or the amplification tube in the nucleic acid amplification kit has a lid that can be opened or closed. In one embodiment of the present invention, the nucleic acid amplification module further comprises a flip mechanism to close and cover the cover of the amplification tube in the nucleic acid amplification module nucleic acid amplification kit, so as to prevent evaporation of liquid and generation of bubbles during amplification from affecting the reaction accuracy.
In the exemplary flip mechanism 42 shown in fig. 9, the flip mechanism has a capping component 421 and a moving component 422. The capping assembly 421 includes a capping plate 423 and a stepping motor 424, and the stepping motor controls the capping plate to move up and down. The motion assembly can move back and forth in a horizontal direction, and the amplification compartment of the nucleic acid amplification module is connected with the motion assembly. When the amplification tube in the amplification compartment or the nucleic acid amplification kit moves to be close to the gland cover plate 423 along with the moving component 422, the stepping motor 424 drives the gland cover plate 423 to move upwards, the cover of the amplification tube can be slowly lifted in the upward movement process, the tube cover is turned over, and the function of turning the cover is realized. When the cover of the amplification tube is lifted to a proper angle, the gland cover plate 423 does not move upwards any more, the moving component 422 continues to move towards the gland cover plate 423, when the amplification tube moves to the position right below the gland cover plate 423, the tube cover is also positioned above the tube orifice, and then the stepping motor 424 drives the cover pressing plate 423 to move downwards to press the tube cover tightly, so that the cover function is realized.
According to the utility model discloses, the liquid distribution module gets the distribution pipe, makes its fluid intercommunication to the distribution head, then moves to elution unit top and absorbs the eluant, then moves to the amplification pipe and discharges the eluant in one or more amplification pipes. Alternatively, the liquid distribution module returns to the elution unit after discharging the eluent, re-draws the eluent and transfers the eluent into another one or more amplification tubes, and so on for one or more cycles. In one embodiment of the present invention, the MIX reagent is placed at the reagent level, which is maintained at 4-8 ℃ during the operation of the apparatus. After the eluent is transferred into the amplification tube, the liquid distribution module transfers the MIX reagent into the amplification tube with the eluent, and the MIX reagent is repeatedly sucked and discharged and uniformly mixed with the reagent system. Thereafter the liquid dispensing module can be moved to the waste zone, removing the dispensing tube, so that a new dispensing tube is used each time MIX reagents are pipetted to the amplification tubes.
Finally, the temperature controller independently heats or cools each amplification tube in a predetermined program, and performs a certain number of thermal cycles to amplify the target nucleic acid. To improve efficiency, each heating and cooling is as rapid as possible. In one embodiment of the present invention, each thermostat also has an associated temperature sensor.
Detection module
The utility model discloses a device is used for the nucleic acid to the sample to amplify and the method that detects, the utility model discloses a device includes detection module 5, and it is used for detecting the nucleic acid of amplification.
The detection module of the present invention is a module suitable for detecting in any way the identifiable label carried by a nucleic acid, including but not limited to fluorescence or other forms of luminescence (e.g., chemiluminescence, bioluminescence, radioluminescence, electroluminescence, electrochemiluminescence, mechanoluminescence, crystallography, thermoluminescence, sonoluminescence, phosphorescence and photoluminescence, etc.), enzymatic reactions, radioactivity, etc. In one aspect of the invention, the amplified nucleic acid is detected by detecting a fluorescent signal carried by the nucleic acid.
As shown in fig. 1 and 2, the device of the present invention has a detection module 5. In one embodiment of the present invention, the detection module is a fluorescence analyzer for exciting and detecting fluorescence, which is fixed or movable to the side of the nucleic acid amplification module, for example, to the side of the amplification tube, and the amplification tube is scanned in a horizontal line.
In the configuration and optical path diagram of the exemplary fluorescence analyzer 51 shown in fig. 10, each optical path is composed of an excitation light source 52, an excitation light filter 53, a first convex lens 54, a dichroic mirror 55, a second convex lens 56, a third convex lens 57, an emission light filter 58, and a detector 59. As shown in fig. 10, the excitation light source 52 is an LED, the emitted light passes through the excitation light filter 53 and the first convex lens 54 to become parallel light, the parallel light is reflected to the second convex lens 56 through the dichroic mirror 55, and is converged and irradiated to a PCR reaction tube, such as an amplification tube 431, so as to excite the sample to emit fluorescence; the excited fluorescence is converted into parallel light by the second convex lens 56, and the parallel light passes through the dichroic mirror 55, and is converged on the detector 59 by the third convex lens 57 and the emission light filter 58, and the detector converts the parallel light into a fluorescence signal. The fluorescence analyzer in the device of the utility model can detect a plurality of fluorescence including FAM, ROX, CY5, VIC and the like. The light path of the fluorescence analyzer can be one light path or a plurality of light paths corresponding to each amplification tube.
The LED light excites the fluorescent molecules in the amplification tube (initially attached to the nucleic acid probes) causing it to fluoresce. At the outset, this fluorescence will typically be effectively blocked by closely spaced quencher molecules. If the target sequence (DNA sequence) of the probe is present in the sample chamber, DNA amplification will occur upon DNA amplification by the TAQ enzyme, and the nucleic acid probe will bind to the amplified target sequence after annealing, leaving the fluorescent molecule and the quencher molecule separated and unable to quench the fluorescence. Fluorescence occurs when a fluorescent molecule is illuminated by excitation light of a certain wavelength. The emitted light is different from the excitation light. Blue incident light is blocked from the detector by a green separate emission filter. The green incident light is similarly blocked from exiting the detector by a yellow emission filter. The fluorescence is captured and propagates via a path into the focusing lens, through the filter and onto the sensitive photodiode.
Liquid dispensing module
The utility model discloses a method that the device is used for extracting, amplifying and detecting the nucleic acid of sample, it includes sample receiving module, one or more schizolysis module, one or more nucleic acid extraction module, nucleic acid amplification module and detection module, and each module still has operating unit separately, and liquid sample shifts in each module and unit.
As shown in fig. 1 and 2, in one embodiment of the invention, the device of the invention comprises a liquid dispensing module 6 for effecting the transfer of the sample and solution.
In one embodiment of the present invention, the liquid dispensing module comprises one or more sensors; one or more dispensing heads; and a displacement unit providing three-dimensional translational freedom of motion for the dispensing head. In one embodiment of the present invention, the sensor may be, for example, an infrared sensor for detecting the presence of dispensing tubing in the liquid dispensing module. For example, an infrared sensor may have an infrared emitter positioned relative thereto, and the presence of the dispensing tube obstructs a line of sight between the emitter and the sensor, whereby the presence or absence of the dispensing tube may be determined.
In one embodiment of the present invention, the sensor in the device further comprises any sensor that controls the degree of horizontal or vertical movement of the dispensing head during the dispensing head pick-up dispensing tube and fluid dispensing operations. For example, in one embodiment of the present invention, the sensor is a force sensor for controlling the degree of vertical movement of the dispensing head during the dispensing head pick up dispensing tube and fluid dispensing operations. The dispensing head may be mounted such that a force acting upwardly against the dispensing head may be sensed by relative movement between the dispensing head and the force sensor. For example, when the dispensing head is forced against a dispensing tube below it, the upward force is transmitted causing a squeeze against the force sensor. The force sensor is then in communication with a processor or controller that controls the vertical movement of the liquid dispensing module so that when an appropriate signal is received from the force sensor, the processor or controller can send instructions to prevent vertical movement of the liquid dispenser. Alternatively, in another embodiment of the present invention, as an alternative to the force sensor, a potential sensor, a magnetic position sensor, a rotary transformer sensor, a proximity sensor, etc. may be used, all of which are included within the scope of the present invention.
In one embodiment of the present invention, the liquid dispensing module comprises one or more dispensing heads configured to receive a dispensing tube per dispensing head. The dispensing head may further comprise a ring movable relative to the dispensing head and configured to remove a dispensing tube fluidly connected to the dispensing head when moved to the end of the dispensing head. Multiple dispense heads may be in parallel and fluidly connected to the same pump at the same time, such that multiple dispense tubes, when connected to the multiple dispense heads, can draw the same amount of liquid, thereby performing multiple parallel assays simultaneously. The dispensing head may also include a number of valves, such as solenoid valves configured to control the flow of air through the dispensing head.
In one embodiment of the invention, the liquid dispensing module includes a pump fluidly connected to one or more dispensing heads and controllable by the controller of the device. The utility model discloses can use various in the device and be fit for the utility model discloses the pump of purpose, including but not limited to piston pump, plunger pump, diaphragm pump, gear pump, slide plate pump, screw pump etc..
Fig. 11 is a schematic diagram of one exemplary liquid dispensing module of the apparatus of the present invention. The liquid distribution module is a single-arm pipette mechanism and is used for transferring sample diluent, PCR samples, PCR reagent loading and the like. The single arm pipette gun mechanism has a single arm pipette gun 61 and a displacement control mechanism that includes a single arm beam 611, a horizontal motion stepper motor 612, a timing belt 613, a wired guide rail 614, an idler 615, and a vertical motion stepper motor 616. The single-arm pipette mechanism may also have a liquid-receiving baffle 617.
The single-arm pipetting gun 61 is fixedly connected with a synchronous belt 613, and the horizontal movement of the single-arm pipetting gun 61 is realized by controlling a horizontal movement stepping motor 612, the synchronous belt 613 and an idler pulley 615. On the other hand, the single-arm pipetting gun 61 is driven to move up and down by controlling the vertical movement stepping motor 616 and the slide block assembly connecting piece.
FIG. 12 is a sectional view of a single-arm pipette tip 61 of the single-arm pipette mechanism, and the single-arm pipette tip 61 sucks and discharges a solution from a pipette tip 6106 detachably located at the lowermost part of the single-arm pipette tip by a piston structure. The piston structure comprises a piston rod 6101, a piston jacket 6102 and a stepping motor 6103, wherein the stepping motor screw 6104 is formed, the stepping motor 6103 drives the piston rod to move up and down through the stepping motor screw 6104, and the liquid taking and discharging functions of the pipette tip are realized. The single-arm pipette also has a tip withdrawing device 6105.
When the single-arm pipette tip 61 is connected with the pipette tip, the stepping motor 6103 of the single-arm pipette tip 61 drives the piston rod to move up and down through the stepping motor lead screw 6104, so as to realize the liquid taking and discharging functions of the pipette tip. When the pipette head moves downwards for liquid suction, the liquid receiving baffle moves to the lower side of the pipette head, and when the pipette head moves upwards, the liquid receiving baffle moves to the position right below the pipette head to realize the liquid receiving function, so that liquid on the outer wall of the pipette head is prevented from falling down and causing pollution.
The process of withdrawing the pipette tip is that when the stepping motor 6103 controls to move to a certain position below, the pipette tip withdrawing device 6105 connected with the upper part is pushed to remove the pipette tip.
Fig. 13 is a schematic diagram of another exemplary liquid dispensing module of the present invention. The liquid distribution module is a liquid transfer mechanism of a discharging gun, is used for sample liquid transfer, nucleic acid extraction, PCR sample adding and the like, and completes the nucleic acid extraction process through the side wall adsorption of the magnet. The liquid transfer mechanism of the liquid transfer gun comprises a liquid transfer gun assembly and a displacement control mechanism. The pipetting gun assembly 62 includes a plurality of pipetting guns 6201 arranged in parallel, a plurality of pumps 6202 and magnet assemblies 6203 connected to the plurality of pipetting guns through hoses in a one-to-one correspondence. The liquid transferring mechanism of the discharging gun realizes the liquid transferring through a plunger pump. The pumps are respectively connected with the corresponding liquid-transfering guns through hoses, and the plungers of the pump assemblies move up and down to finish liquid suction and discharge. The gun-discharging pipetting mechanism completes the side wall adsorption and release of the magnetic particles and the nucleic acid combined with the magnetic particles through a magnet.
The pipetting gun assembly 62 may also have a pipette tip retracting mechanism comprising a stepper motor 6206 that controls the pipetting gun to move up and down, a pipette tip push plate 6204, and a bayonet 6205. The bayonet 6205 is pushed by an electromagnet and can be pushed out forwards to be clamped above the pipetting gun head push plate 6204, the pipetting gun is enabled to move upwards through the stepping motor 6206, and the pipetting gun head push plate cannot move upwards under the blocking of the bayonet so as to finish the process of withdrawing and transferring the liquid gun head.
The pipetting gun assembly 62 is moved in position by Y-axis, Z-axis motion.
In one embodiment of the present invention, the displacement control mechanism of the pipetting gun assembly 62 is a gantry, such as a gantry having two or three translational degrees of freedom, driven by a coded stepper motor driven drive slide.
When the dispensing head to be controlled is a separate single dispensing head (e.g. a single-arm pipette), it may be mounted on a gantry with three translational degrees of freedom X (horizontal), Y (front-back), Z (up-down), so that it can be moved within the device to other modular units or kits located in the units, e.g. dilution tubes of dilution units, amplification tubes in compartments of nucleic acid amplification modules or containers containing MIX reagents. When a plurality of distribution heads are connected in parallel, it can cover the utility model discloses its whole horizontal range that needs to arrive in the device, consequently can install on the rack that has two translation degrees of freedom of Y (front and back), Z (from top to bottom), and need not the utility model discloses horizontal migration in the device. The parallel dispense heads may simultaneously pick up one or more dispense tubes and then move to a desired location, such as a sample tube, a lysis tube, an elution tube, etc., to draw liquid from one or more tubes and then transfer it to a desired destination location. In an embodiment of the present invention, the stage with two translational degrees of freedom and the stage with three translational degrees of freedom can coexist, which jointly constitute the displacement unit of the present invention.
In one embodiment of the invention, the sensor and the dispensing head are mounted on a displacement unit. The pump may also be mounted on the displacement unit, or on another part of the device, in fluid connection with the dispensing head via a tube. The mounting means may be by mechanical fastening, such as one or more screws.
Reagent and consumable set-up zone
The device of the utility model is used for extracting, amplifying and detecting the nucleic acid of the sample, and comprises various reagents and consumables. As shown in fig. 1 and 2, in one embodiment of the present invention, the device includes reagent and consumable material setting areas, such as pipette tip setting area 7, PCR reagent storage and mixing processing area 8, waste area 9, etc.
In an embodiment of the present invention, the device comprises a pipette tip setting area. The setting area of the liquid-transfering gun head can be in any form, so long as the liquid-transfering gun head can be conveniently accommodated and replaced in the operation process of the device.
In one embodiment of the present invention, the device includes a PCR reagent storage and mixing treatment area. The PCR reaction requires the use of reagents such as enzymes, reaction buffers, and water. These reagents may be stored separately in advance, or may be mixed in advance in a desired ratio. The PCR reagent storage and mixing treatment area can also be provided with a temperature control system for cooling and heat preservation treatment of relevant reagents.
In one embodiment of the present invention, the device includes a waste zone. The waste material district can be any form as long as can hold the utility model discloses the waste material that the device operation in-process produced can. For example, the waste region may be a bucket or bag having an opening that can contain a liquid.
Automated molecular in vitro diagnostic instrument
The utility model also provides an automatic molecular in vitro diagnostic instrument. In one embodiment of the present invention, the molecular in vitro diagnostic apparatus comprises the aforementioned nucleic acid extraction and detection device, and a computer-based automatic control system. The automatic system can control the liquid distribution module of the molecular in-vitro diagnostic instrument to transfer samples between or in the sample receiving module, the cracking module, the nucleic acid extracting module and the nucleic acid amplifying module of the nucleic acid extracting and detecting device in a programmed manner, and perform cracking, extracting, amplifying, detecting and other procedures according to preset steps under proper conditions to detect the samples to be detected. The diagnostic instrument can be used for identifying infection sources, detecting genetic diseases, detecting cancers or detecting genetic variation.
In one embodiment of the invention, the diagnostic instrument can be used for the detection of: influenza virus, enterovirus, hepatitis B virus, hepatitis C virus, Ebola virus, Marburg virus, SARS virus, Securia virus, bunyavirus, rhinovirus, respiratory nucleocapsid virus, cholera virus, etc., or bacterial pathogens such as tubercle bacillus, Escherichia coli, Acinetobacter baumannii, Diplococcus pneumoniae, Streptococcus lactis, Sporosarcina urens, Staphylococcus aureus, Bacillus subtilis, Bacillus anthracis, Bacillus subtilis, Bacillus streptococci, Bacillus proteus, Vibrio cholerae, Treponema pallidum, etc.
In one embodiment of the invention, the diagnostic instrument may be used for the detection of: gastric cancer, liver cancer, lung cancer, esophageal cancer, cervical cancer, breast cancer, colon cancer, rectal cancer, nasopharyngeal cancer, ovarian cancer, renal cancer, bladder cancer, thyroid cancer, skin cancer, etc.; malignant tumors derived from mesenchymal tissues such as muscle, fat, bone, blood vessel, lymph, etc., for example, rhabdomyosarcoma, leiomyosarcoma, fibrosarcoma, liposarcoma, osteosarcoma, chondrosarcoma, angiosarcoma, lymphosarcoma, etc. Also, for example, leukemia, Hodgkin's disease, Wilms ' tumor (nephroblastoma), melanoma, retinoblastoma, seminoma, granuloma, Kupffer's tumor, Ewing's tumor, malignant vascular endothelial cell tumor, or Paget's disease of the breast. Analysis and diagnosis.
Although the present invention has been described in detail with respect to the exemplary embodiments and advantages thereof, it should be understood that various changes, substitutions, and alterations can be made hereto without departing from the spirit and scope of the invention as defined by the appended claims.
Claims (28)
1. A nucleic acid extraction and detection device, the device comprising:
one or more sample receiving modules, each sample receiving module having a compartment adapted to receive a sample or sample container;
one or more lysis modules, each lysis module having a compartment adapted to receive a lysis reagent or a lysis kit;
one or more nucleic acid extraction modules, each nucleic acid extraction module having a compartment adapted to receive a nucleic acid extraction reagent or nucleic acid extraction kit;
one or more nucleic acid amplification modules, each nucleic acid amplification module having a plurality of compartments adapted to receive nucleic acid amplification reagents or nucleic acid amplification kits; and
a detection module for detecting the position of the optical fiber,
characterized in that the nucleic acid extraction module has one or more nucleic acid extraction units consisting of a binding unit, a washing unit and an elution unit, and in that the nucleic acid amplification module has one or more nucleic acid amplification units, the nucleic acid amplification unit corresponding to the nucleic acid extraction sample obtained by each nucleic acid extraction unit in the nucleic acid extraction module comprising compartments for performing two or more amplification reactions.
2. The apparatus of claim 1, wherein said sample receiving module has a sample holder for receiving a sample tube and is disposed at a front end of said apparatus adjacent to the operator.
3. The device of claim 1, wherein the sample is lysed in the lysis module using a lysing agent, an enzyme, ultrasound, or physical abrasion.
4. The device of claim 1, wherein the lysis module further comprises means for lysing the sample.
5. The device of claim 4, wherein the means for lysing the sample is an oscillator comprising an eccentric shaft, an eccentric holder, and a motor, the eccentric shaft rotating about the axis of the motor to oscillate the eccentric holder.
6. The device of claim 3, wherein the lysis module is adapted to receive abrasive particles or a lysis kit comprising abrasive particles to lyse the sample by physical abrasion.
7. The device of claim 6, wherein the lysis kit has a lysis tube that can contain abrasive particles.
8. The apparatus of claim 7, wherein the lysis tube has a cap and the inner wall has two or more axial strips.
9. The device of claim 3, wherein the lysis module is adapted to receive a lysis agent or a lysis kit comprising a lysis agent to chemically or biologically lyse the sample.
10. The device of claim 1, wherein the nucleic acid extraction module has a magnet device that can apply a magnetic field, control a magnetic material and/or the magnetic material that binds nucleic acids.
11. The device of claim 10, which has a dispensing tube insertable into the binding unit, the washing unit and the elution unit, wherein the nucleic acid in the container is extracted by a magnetic bead method, wherein after the nucleic acid sample is bound to the magnetic beads in the solution in the container, the tip of the dispensing tube is inserted into the bottom of the container, the magnet is close to the dispensing tube, the magnetic beads and the bound sample are attracted to the side wall of the dispensing tube by the magnet, the dispensing tube moves to another container after the liquid is discharged, the magnet leaves the dispensing tube, the magnetic field is removed, and the attracted magnetic beads and the sample are released.
12. The apparatus of claim 1, wherein the nucleic acid amplification module is configured to perform isothermal amplification or PCR.
13. The apparatus of claim 12, wherein the nucleic acid amplification module is configured to perform fluorescent quantitative PCR of DNA and RNA.
14. The device of claim 1, wherein each nucleic acid amplification unit is capable of performing two or more identical or different amplification reactions on the same nucleic acid extraction sample.
15. The apparatus of claim 1, wherein the nucleic acid amplification module further comprises a temperature regulator that independently heats or cools each of the plurality of compartments such that each compartment independently performs a different or the same amplification reaction.
16. The apparatus of claim 1, wherein the nucleic acid amplification module further comprises a flip mechanism to close the lid of the amplification tube.
17. The apparatus of claim 16, wherein the flip mechanism has a capping component and a moving component.
18. The apparatus of claim 17, wherein the gland assembly includes a gland plate and a stepper motor, the stepper motor controlling the gland plate to move up and down; the motion assembly can move back and forth in the horizontal direction, and the amplification compartment of the nucleic acid amplification module is connected with the motion assembly; the cover turning mechanism is arranged in a way that when an amplification tube in an amplification compartment or a nucleic acid amplification kit moves to be close to the cover pressing plate along with the movement component, the stepping motor drives the cover pressing plate to move upwards, the cover of the amplification tube is slowly lifted, the tube cover is turned over, when the cover of the amplification tube is lifted to a proper angle, the cover pressing plate does not move upwards any more, the movement component continues to move towards the cover pressing plate, and then the stepping motor drives the cover pressing plate to move downwards, so that the tube cover is pressed.
19. The device of claim 1, wherein the detection module detects an identifiable tag carried by the nucleic acid.
20. The device of claim 19, wherein said identifiable marker is luminescent, enzymatic, radioactive.
21. The device of claim 20, wherein said identifiable marker is fluorescent.
22. The device of claim 1, wherein the detection module comprises a fluorescence analyzer.
23. The device of claim 1, further comprising a liquid distribution module to transfer or distribute liquid in the device between two or more locations.
24. The apparatus of claim 23, wherein the liquid dispensing module comprises: one or more sensors; one or more dispensing heads; and a displacement control mechanism providing three-dimensional translational motion freedom for the dispensing head.
25. The device of claim 1, further comprising reagent and consumable set-up zones.
26. The apparatus of claim 25, wherein the reagent and consumable set-up zones comprise one or more of a pipette tip set-up zone, a PCR reagent storage and mixing treatment zone, and a waste zone.
27. An automated molecular in vitro diagnostic apparatus comprising the nucleic acid extraction and detection device of any one of claims 1 to 26, and an automated control system.
28. The molecular in vitro diagnostic apparatus according to claim 27, characterized in that said automatic control system is a computer-based automatic control system.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN112553042A (en) * | 2019-09-10 | 2021-03-26 | 嘉兴市艾科诺生物科技有限公司 | Apparatus and method for nucleic acid extraction and detection |
| CN113583803A (en) * | 2021-07-26 | 2021-11-02 | 上海思路迪生物医学科技有限公司 | Nucleic acid extraction and amplification POCT consumable and method applied to molecular diagnosis |
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| CN113174328B (en) * | 2021-06-15 | 2024-05-03 | 中国科学院苏州生物医学工程技术研究所 | Uncapping module and nucleic acid detection device comprising same |
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| CN113652343A (en) * | 2021-07-30 | 2021-11-16 | 北京威妙生物科技有限公司 | Multi-project full-automatic molecular detector |
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| CN112553042A (en) * | 2019-09-10 | 2021-03-26 | 嘉兴市艾科诺生物科技有限公司 | Apparatus and method for nucleic acid extraction and detection |
| CN113583803A (en) * | 2021-07-26 | 2021-11-02 | 上海思路迪生物医学科技有限公司 | Nucleic acid extraction and amplification POCT consumable and method applied to molecular diagnosis |
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