CN111849758A - Full-automatic real-time fluorescent quantitative PCR detection system - Google Patents
Full-automatic real-time fluorescent quantitative PCR detection system Download PDFInfo
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- CN111849758A CN111849758A CN202010828644.5A CN202010828644A CN111849758A CN 111849758 A CN111849758 A CN 111849758A CN 202010828644 A CN202010828644 A CN 202010828644A CN 111849758 A CN111849758 A CN 111849758A
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- C—CHEMISTRY; METALLURGY
- 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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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L7/00—Heating or cooling apparatus; Heat insulating devices
- B01L7/52—Heating or cooling apparatus; Heat insulating devices with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples
Abstract
The invention discloses a full-automatic real-time fluorescent quantitative PCR detection system, which comprises a detection main body, wherein the detection main body consists of a shell component and an inner component, the shell component consists of a shell, a screen and a bin gate, and the inner component consists of an electric control component, a magnetism shifting component, a kit loading and unloading component, a core module, an XY axis moving component, a liquid transferring component, a screw cap component, an air duct and an optical component; the software architecture of the detection main body comprises the following components: the system comprises a UI layer, a data control layer and a data link layer, wherein the data control layer is divided into a data management, a logic control and algorithm library, and the data link layer comprises a configuration file, a database, a communication protocol and a physical interface. The invention has reasonable structure, integrates two functions of automatic nucleic acid extraction and automatic PCR amplification, and realizes automatic detection process.
Description
Technical Field
The invention relates to the technical field of gene and life science instruments, in particular to a full-automatic real-time fluorescence quantitative PCR detection system.
Background
The nucleic acid detection aims at virus detection of clinical diseases such as virus DNA and RNA, hepatitis B, hepatitis C, AIDS and the like, and mainly detects whether virus nucleic acid exists in blood or not as a basis for diagnosing whether pathogen infection exists in an organism or not.
The existing nucleic acid detection process is low in automation degree, needs to perform professional technical culture on operators, has certain biological hazards to the operators, particularly needs to consume a large amount of time for some high-risk samples, and also has certain potential safety hazards.
Disclosure of Invention
The invention aims to solve the defects in the prior art, and provides a full-automatic real-time fluorescent quantitative PCR detection system which integrates two functions of automatic nucleic acid extraction and automatic PCR amplification and realizes an automatic detection process.
In order to achieve the purpose, the invention adopts the following technical scheme:
the full-automatic real-time fluorescent quantitative PCR detection system comprises a detection main body, wherein the detection main body consists of a shell component and an inner component, the shell component consists of a shell, a screen and a bin door, and the inner component consists of an electric control component, a magnetism shifting component, a kit loading and unloading component, a core module, an XY axis moving component, a liquid transferring component, a screw cover component, an air duct and an optical component;
the software architecture of the detection main body comprises the following components: the system comprises a UI layer, a data control layer and a data link layer, wherein the data control layer is divided into a data management, logic control and algorithm library, and the data link layer comprises a configuration file, a database, a communication protocol and a physical interface;
the system software of the detection main body comprises a user interaction system, a detection control system and a detection result processing system, wherein the user interaction system comprises a detection module, a review module, a setting module and a maintenance module, and the detection control system comprises an XY level control module, a liquid transfer device control module, an optical control module, a temperature control module, a rotary cover control module and a dial rod control module.
Preferably, the user interaction system is used for daily operation control, and the detection module comprises detection guide, sample information entry, detection information confirmation, detection monitoring, detection result display and detection result curve display.
Preferably, the review module comprises a review list, a condition query, a detection result display and a detection result curve display.
Preferably, the setting module includes time setting, network setting and hardware self-checking, and the maintenance module includes software upgrading, radio frequency card information, factory restoration setting and log export.
Preferably, the XY level control module controls a consumable level, a reagent level, a purification level, a waste liquid level, a PCR preparation level, and a PCR amplification level.
Preferably, pipettor control module control loading and unloading consumptive material, reagent pipe thorn membrane, liquid suction row and liquid mixing, spiral cover control module control PCR tube cap, the driving lever control module is the electro-magnet.
Preferably, the optical control module controls 4-channel LED control and 4-channel optical acquisition, and the temperature control module is used for purification bin temperature acquisition control, PCR temperature acquisition control and radiator temperature acquisition control.
Preferably, the detection result processing system performs crosstalk correction on the 4-channel fluorescence data, performs baseline correction and calculates a CT value, and finally gives result judgment and curve display according to the CT value of the 4 th channel internal parameter.
Compared with the prior art, the invention has the beneficial effects that:
according to the invention, only a sample needs to be manually added, the nucleic acid extraction is automated, and after the PCR amplification is automatically carried out, an experimental result and a result curve are displayed, so that the professional skill and time required by manual operation are greatly reduced; the biological harm of the sample to operators is reduced, particularly for some inactivated high-risk samples, the time for the operators to process and extract the sample is really saved, and the high integration of safe, convenient and fast full-automatic nucleic acid extraction and full-automatic PCR amplification is really realized.
Drawings
FIG. 1 is a schematic diagram of the internal components of the full-automatic real-time fluorescence quantitative PCR detection system according to the present invention;
FIG. 2 is a schematic view of the housing assembly of the fully automatic real-time fluorescence quantitative PCR detection system according to the present invention;
FIG. 3 is a schematic diagram of a software architecture of the full-automatic real-time fluorescence quantitative PCR detection system according to the present invention;
FIG. 4 is a diagram of a user interaction system of the fully automatic real-time fluorescence quantitative PCR detection system according to the present invention;
FIG. 5 is a schematic diagram of a control software architecture of the full-automatic real-time fluorescence quantitative PCR detection system according to the present invention;
FIG. 6 is a flow chart of the result processing of the full-automatic real-time fluorescence quantitative PCR detection system according to the present invention.
In the figure: the device comprises an electric control assembly 1, a magnetic poking assembly 2, a kit assembling and disassembling assembly 3, a core module 4, an XY axis moving assembly 5, a liquid transfer assembly 6, a screw cover assembly 7, an air duct 8, an optical assembly 9, a screen 10, a shell 11 and a bin door 12.
Detailed Description
In order to make the aforementioned objects, features and advantages of the present invention comprehensible, embodiments accompanied with figures are described in detail below. In the following description, numerous specific details are set forth in order to provide a thorough understanding of the present invention. This invention may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein, but rather should be construed as broadly as the present invention is capable of modification in various respects, all without departing from the spirit and scope of the present invention.
It will be understood that when an element is referred to as being "secured to" another element, it can be directly on the other element or intervening elements may also be present. When an element is referred to as being "connected" to another element, it can be directly connected to the other element or intervening elements may also be present. The terms "vertical," "horizontal," "left," "right," and the like as used herein are for illustrative purposes only and do not represent the only embodiments.
Referring to fig. 1-6, the full-automatic real-time fluorescence quantitative PCR detection system comprises a detection main body, wherein the detection main body comprises a shell component and an inner component, the shell component comprises a shell 11, a screen 10 and a bin door 12, and the inner component comprises an electric control component 1, a magnetism shifting component 2, a kit assembling and disassembling component 3, a core module 4, an XY axis moving component 5, a liquid transferring component 6, a screw cap component 7, an air duct 8 and an optical component 9;
the software architecture of the detection subject is divided into: a UI layer, a data control layer and a data link layer, wherein the UI layer is mainly responsible for displaying a UI interface, and UI logics such as interface jumping, interface closing and the like, the data control layer is divided into a data management, logic control and algorithm library, the data management is system data, UI data and a unified manager of communication data, the logic control controls system data storage logic, UI data display updating logic and communication data updating and storage logic, the data link layer comprises a configuration file, a database, a communication protocol and a physical interface, the configuration file stores a series of configuration related files such as system configuration, project configuration and the like, the database stores related data generated in measurement, the communication protocol provides encapsulation and analysis of the protocol, obtaining related protocol command data through a communication protocol class, wherein a physical interface provides a related communication algorithm for communicating with peripheral equipment and mainly comprises sending and reading of data;
the system software of the detection main body consists of three parts, namely a user interaction system, a detection control system and a detection result processing system, wherein the user interaction system comprises a detection module, a review module, a setting module and a maintenance module, the detection control system comprises an XY level control module, a pipettor control module, an optical control module and a temperature control module, spiral cover control module and dial rod control module, XY horizontal control module control consumptive material position, the reagent position, the purification position in storehouse, the waste liquid position in storehouse, PCR preparation position and PCR amplification position, pipettor control module control loading and unloading consumptive material, reagent piping thorn membrane, liquid suction row and liquid mixing, spiral cover control module control PCR tube cap, dial rod control module is the electro-magnet, 4 passageway LED controls and 4 passageway optical acquisition are controlled to optical control module, temperature control module is used for purification storehouse temperature acquisition control, PCR temperature acquisition control and radiator temperature acquisition control.
The control system mainly transmits and receives control instructions through a CAN bus, and each unit module is responsible for executing corresponding control instructions and returning response instructions, execution results or error codes; wherein: the states and the reset of all modules of the instrument are detected through an optical coupler, and the states of all unit modules are judged through signal input and output of an I/O port.
The user interaction system is used for daily operation control, the detection module comprises a detection guide, sample information input, detection information confirmation, detection monitoring, detection result display and detection result curve display, the review module comprises a review list, condition query, detection result display and detection result curve display, the setting module comprises time setting, network setting and hardware self-checking, and the maintenance module comprises software upgrading, radio frequency card information, factory setting recovery and log export.
And the detection result processing system performs crosstalk correction on the 4-channel fluorescence data, performs baseline correction and calculates a CT value, and finally gives result judgment and curve display according to the CT value of the 4 th channel internal parameter.
The principle of nucleic acid extraction: the kit is put into an instrument, and an RFID card is input into an experiment control program and an interpretation index. And during the vertical movement of the mechanical arm with the pipette head at different positions of the purifying cabin, continuously sucking and discharging liquid to uniformly mix the liquid and the magnetic beads. The magnet module is stirred by a manipulator to carry out magnetic attraction.
Cracking: in the cracking process, a mechanical arm with a pipette tip is continuously and uniformly mixed with cracking solution, a sample and magnetic beads, pathogen cells in the sample are cracked by heating through a heating module, nucleic acid in cell nucleuses is released, PK enzyme degrades protein, and nucleic acid and internal reference are adsorbed on the surfaces of the magnetic beads. After the cracking is finished, the magnet module is shifted to the magnetic attraction position through the manipulator to carry out magnetic attraction, and the magnetic beads with the adsorbed nucleic acids are magnetically attracted on one side. After the magnetism is inhaled, the arm of taking the liquid-transfering gun head constantly shifts the waste liquid of opposite side to the waste liquid cabin. Finally, the magnetic suction module is reset again through the mechanical arm;
washing: and the mechanical arm with the pipette head transfers the cleaning solution to the purification cabin. During the up-and-down movement of the mechanical arm with the pipette head at different positions of the purification cabin, the washing liquid and the magnetic beads are uniformly mixed by continuous liquid suction and discharge, so that more impurities are dissolved in the liquid. After fully mixing, carry out magnetism once more and inhale, the arm of taking the liquid-transfering gun head shifts the waste liquid to the waste liquid cabin. Finally, the magnetic suction module is reset again through the mechanical arm;
and (3) elution: the mechanical arm with pipette tip transfers the eluent to the purification cabin. During the up-and-down motion of the mechanical arm with the pipette head at different positions of the purification cabin, the eluent and the magnetic beads are uniformly mixed through continuous liquid suction and drainage, the nucleic acid adsorbed on the surfaces of the magnetic beads is separated from the liquid through heating of the heating module, the magnetic beads are adsorbed on one side after the nucleic acid is sufficiently and uniformly mixed, and the purified nucleic acid is finally obtained in the liquid.
The preparation principle is as follows: preparation: the mechanical arm loosens the tube cover of the PCR tube on the kit, the mechanical arm with the pipette head adds the product into the PCR dry powder tube, and the mechanical arm with the pipette head continuously sucks and discharges the liquid up and down to uniformly mix the product with the PCR dry powder reagent. Then, paraffin oil was added as a seal to the upper layer of the liquid in the PCR tube by a mechanical arm with a pipette tip. And finally, screwing the PCR tube cap by the mechanical arm to finish the preparation.
PCR amplification principle: after the automatic preparation of the product is finished, specific fragments and internal references of nucleic acid in a sample participate in reaction with raw materials such as primers, probes, enzymes, dNTP and the like in a PCR reagent, and a heating module and a heat dissipation module execute PCR amplification tribasic: denaturation, annealing and extension. And after each cycle is finished, the XY motion module moves to the optical acquisition module to acquire a fluorescence value for one time until the PCR cycle is finished. Finally, after the experiment is finished, the software automatically judges whether the test card is positive or negative according to the judgment indexes in the RFID card, and displays the corresponding CT value and the result curve on a user interface.
The device is generally applied to clinical molecular detection laboratories of medical institutions, detection departments of hospitals and other institutions and detection laboratories. This device does not support mobile in-vehicle applications and closed environment use.
The instrument has certain requirements on the use environment and can meet the following requirements:
a) working temperature: 10-30 deg.C
b) Relative humidity: 20 to 85 percent
c) Working voltage: the frequency of the AC 220V, 50Hz,
d) rated power: 220VA
e) Atmospheric pressure: 86.0KPa to 106.0KPa
The above description is only for the preferred embodiment of the present invention, but the scope of the present invention is not limited thereto, and any person skilled in the art should be considered to be within the technical scope of the present invention, and the technical solutions and the inventive concepts thereof according to the present invention should be equivalent or changed within the scope of the present invention.
Claims (8)
1. The full-automatic real-time fluorescence quantitative PCR detection system comprises a detection main body and is characterized in that the detection main body consists of a shell component and an inner component, the shell component consists of a shell (11), a screen (10) and a bin door (12), and the inner component consists of an electric control component (1), a magnetism shifting component (2), a kit assembling and disassembling component (3), a core module (4), an XY axis moving component (5), a liquid shifting component (6), a screw cover component (7), an air duct (8) and an optical component (9);
the software architecture of the detection main body comprises the following components: the system comprises a UI layer, a data control layer and a data link layer, wherein the data control layer is divided into a data management, logic control and algorithm library, and the data link layer comprises a configuration file, a database, a communication protocol and a physical interface;
the system software of the detection main body comprises a user interaction system, a detection control system and a detection result processing system, wherein the user interaction system comprises a detection module, a review module, a setting module and a maintenance module, and the detection control system comprises an XY level control module, a liquid transfer device control module, an optical control module, a temperature control module, a rotary cover control module and a dial rod control module.
2. The fully automatic real-time quantitative fluorescence PCR detection system of claim 1, wherein the user interaction system is used for daily operation control, and the detection module comprises detection guide, sample information entry, detection information confirmation, detection monitoring, detection result display and detection result curve display.
3. The fully automated real-time quantitative fluorescence PCR detection system of claim 1, wherein the review module comprises a review list, a conditional query, a detection result display, and a detection result curve display.
4. The full-automatic real-time fluorescent quantitative PCR detection system according to claim 1, wherein the setting module comprises time setting, network setting and hardware self-checking, and the maintenance module comprises software upgrading, radio frequency card information, factory setting restoration and log export.
5. The full-automatic real-time fluorescent quantitative PCR detection system of claim 1, wherein the XY level control module controls a consumable site, a reagent site, a purification site, a waste solution site, a PCR preparation site, and a PCR amplification site.
6. The full-automatic real-time fluorescent quantitative PCR detection system according to claim 1, wherein the pipettor control module controls handling consumables, a reagent tube piercing film, liquid suction and discharge and liquid mixing, the screw cap control module controls a PCR tube cap, and the dial rod control module is an electromagnet.
7. The full-automatic real-time fluorescent quantitative PCR detection system of claim 1, wherein the optical control module controls 4-channel LED control and 4-channel optical collection, and the temperature control module is used for purification bin temperature collection control, PCR temperature collection control and radiator temperature collection control.
8. The full-automatic real-time fluorescence quantitative PCR detection system of claim 1, wherein the detection result processing system performs crosstalk correction on 4-channel fluorescence data, performs baseline correction and calculates CT values, and finally gives result judgment and curve display according to the CT value of the 4 th channel internal parameter.
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Cited By (1)
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