CN202415561U - Quantitative PCR (Polymerase Chain Reaction) microfluidic control chip device - Google Patents
Quantitative PCR (Polymerase Chain Reaction) microfluidic control chip device Download PDFInfo
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- CN202415561U CN202415561U CN 201220018703 CN201220018703U CN202415561U CN 202415561 U CN202415561 U CN 202415561U CN 201220018703 CN201220018703 CN 201220018703 CN 201220018703 U CN201220018703 U CN 201220018703U CN 202415561 U CN202415561 U CN 202415561U
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Abstract
The utility model discloses a quantitative PCR (Polymerase Chain Reaction) microfluidic control chip device, which comprises a fixed base, three independent thermostats, a PCR microfluidic control chip and an electrochemical detector and is characterized in that the three independent thermostats are arranged in the fixed base; the three independent thermostats are a low-temperature annealing thermostat, a favorable-temperature extension thermostat and a high-temperature denaturation thermostat; the low-temperature annealing thermostat, the favorable-temperature extension thermostat and the high-temperature denaturation thermostat are arranged in sequence side by side; PCR reaction in the PCR microfluidic control chip can be finished in a low-temperature annealing region, a favorable-temperature extension region and a high-temperature denaturation region; and a DNA (Deoxyribonucleic acid) amplification reaction tube at the favorable-temperature extension region is internally provided with an electrochemical detector for quantitative detection of a PCB reactant. The quantitative PCR microfluidic control chip is applicable to various gene diagnosis detection and laboratory use. The quantitative PC microfluidic control chip device integrates temperature control, PCR reaction and detection into a whole and has high microminiaturization degree.
Description
Technical field
The utility model belongs to a kind of quantitative PCR micro flow control chip device; Specifically; Be that electrochemical measuring technique and round pcr are integrated in the micro-fluidic chip system; Bring into play integrated, multi-functional advantage, this micro-fluidic chip is applicable to range gene diagnostic detection and laboratory use, belongs to technical field of biological.
Background technology
Polymerase chain reaction (English full name: Polymerase Chain Reaction, be called for short PCR) be a kind of method of the synthetic specific DNA fragment of external enzymatic have high specificity, highly sensitive, easy and simple to handle, characteristics such as save time.PCR extends three primitive reaction steps by high-temperature denatured, low-temperature annealing (renaturation) and thermophilic and forms one-period, repeatedly circulates.Circulation of every completion only needs about several minutes, just can be able to increase rapidly to amplify be about 10 by goal gene expanded in about 2-4 hour
9Individual copy, it not only can be used for fundamental researchs such as gene isolation, clone and nucleic acid sequence analysis, also can be used for the diagnosis of major diseases such as genetic diseases, tumour.Round pcr is revolutionary invention and the milestone in the biomedical sector.
The micro-fluidic chip technology is one of research forward position focus of present analysis science.The micro-fluidic chip technology is with laboratory The whole analytical process such as sample preparation, sample filtering, sample separation so that sample detection etc. are integrated in carries out on the little chip block, can realize the robotization and the microminiaturization of The whole analytical process.
An important focus is studied PCR integrated technology in the micro-fluidic chip exactly in the micro-fluidic chip field.At present, integrated PCR micro-fluidic chip technology generally is only to design round pcr in micro-fluidic chip, to accomplish, and the PCR micro-fluidic chip is a prior art, but it lacks real time quantitative measurement technology, has limited the range of application of integrated PCR micro-fluidic chip.Therefore, realize the integrated of quantitative measurement technology and PCR micro-fluidic chip, the applied research field that can further widen the PCR micro-fluidic chip, so electrophoresis and dyeing course when this method has been avoided the PCR product analysis simultaneously are more quick, easy and sensitive.
Summary of the invention
The utility model purpose is to propose a kind of low preparation cost, quantitative PCR micro flow control chip device easy and simple to handle.
The technical scheme that the utility model adopted is: the utility model is described
The quantitative PCR micro flow control chip deviceComprise independently thermostatted of firm banking, three; PCR micro-fluidic chip and electrochemical detector; It is characterized in that on firm banking, being manufactured with three independently thermostatteds; Three independently thermostatted is respectively the low-temperature annealing thermostatted, thermophilic extends thermostatted and high-temperature denatured thermostatted; Described low-temperature annealing thermostatted, thermophilic extend thermostatted and high-temperature denatured thermostatted is arranged side by side in regular turn; Described low-temperature annealing thermostatted, thermophilic extend on thermostatted and the high-temperature denatured thermostatted and are formed with accurately three humidity provinces in low-temperature annealing district, thermophilic extension area and the high-temperature denatured district of controlled temperature respectively, in the low-temperature annealing district, thermophilic extension area and high-temperature denatured district have three required temperature of basic PC R reactions step respectively, above-mentioned three independently thermostatted above be placed with the PCR micro-fluidic chip; In the PCR micro-fluidic chip, be distributed with the amplified reaction pipe; The amplified reaction pipe has PCR reaction fluid inlet, PCR reaction amplification back liquid outlet and dna amplification reaction pipe, and PCR reaction fluid inlet is positioned at high-temperature denatured district and is connected with miniature syringe pump, and the amplified reaction pipe passes high-temperature denatured district, thermophilic extension area and low-temperature annealing district successively; The amplified reaction pipe is as the pcr amplification reaction district in the PCR micro-fluidic chip, and the PCR reaction in the PCR micro-fluidic chip can be accomplished in low-temperature annealing district, thermophilic extension area and high-temperature denatured district; In the dna amplification reaction pipe of thermophilic extension area, be provided with the electrochemical detector that the detection by quantitative of PCR reactant is used.PCR annealing region integrated electrochemical detecting electrode in the micro-fluidic chip is used for the detection of goal gene number of amplification.
Described quantitative PCR micro flow control chip device, its characteristics be described three independently thermostatted be the miniature rectangular thermostatted of processing by copper, aluminium or stainless material.
Described quantitative PCR micro flow control chip device, its characteristics are that described electrochemical detector comprises working electrode, counter electrode and reference electrode.
The device of the utility model can pass through this independent temperature heating unit of thermostatted; Accurately control unwind, the temperature of annealing and elongating temperature; Control time and the loop cycle number of times of PCR reaction solution through microchannel length and rate of flow of fluid in the micro-fluidic chip in each humidity province; The amplification of realization goal gene; Utilize electrochemical measuring technique detection by quantitative goal gene number of amplification at each cyclic annealing step place, more more quick, easy, sensitive than regular-PCR and fluorescent PCR, electrophoresis and dyeing course when this method has been avoided the PCR product analysis simultaneously.
The characteristics of the utility model: three of the utility model micro constant-temperature devices independently, can accurately control the temperature that high-temperature denatured, low-temperature annealing (renaturation) and thermophilic extend three primitive reaction steps, realize the PCR reaction in the micro-fluidic chip.PCR annealing region integrated electrochemical detecting electrode in the micro-fluidic chip; Can be to goal gene number of amplification detection by quantitative; Detection time, cost, the dye discoloration of experiment have been improved greatly; Electrophoresis and dyeing course when having avoided the PCR product analysis have been widened the research range of PCR integrated microfluidic chip.This apparatus platform becomes one temperature control, pcr amplification, detection three segment sets, and is quick, easy, and integrated degree is high.The utility model the quantitative PCR micro flow control chip device be applicable to that range gene diagnostic detection and laboratory use.Can further widen the applied research field of PCR micro-fluidic chip, so electrophoresis and dyeing course when this method has been avoided the PCR product analysis simultaneously are more quick, easy and sensitive.This apparatus platform becomes one temperature control, PCR reaction, detection three segment sets, and microminiaturized degree is high, and this device making method very easily is used more widely.
Description of drawings
Fig. 1 is the quantitative PCR micro flow control chip device schematic top plan view of the utility model.
Fig. 1-1 is the A part partial enlarged drawing of Fig. 1.
Fig. 2 is the quantitative PCR micro flow control chip device schematic cross-section of the utility model.
Fig. 3 is that the reacted detected result figure of PCR takes place on this device the signal indicator.
Among the figure: 1, the low-temperature annealing district; 2, the thermophilic extension area; 3, high-temperature denatured district; 4, PCR reacts fluid inlet; 5, the dna amplification reaction district; 6, the PCR micro-fluidic chip; 7, firm banking; 8, electrochemical detector; 9, PCR reaction amplification back liquid outlet; 10, working electrode; 11, counter electrode; 12, reference electrode; 13, detection signal curve in the time of before the PCR reaction expansion; 14, PCR reaction expansion back detection signal curve.
Embodiment
Right below in conjunction with embodiment and accompanying drawingThe utility model
Be elaborated:
Shown in Fig. 1, Fig. 1-1 and 2; The quantitative PCR micro flow control chip device that the utility model provided; Comprise independently thermostatted of firm banking 7, three; PCR micro-fluidic chip and electrochemical detector; Described thermostatted, PCR micro-fluidic chip and electrochemical detector are prior art; It is characterized in that on firm banking 7, being manufactured with three independently thermostatteds, three independently thermostatted is respectively the low-temperature annealing thermostatted, thermophilic extends thermostatted and high-temperature denatured thermostatted, and described low-temperature annealing thermostatted, thermophilic extend thermostatted and high-temperature denatured thermostatted is arranged side by side in regular turn; Be formed with accurately three humidity provinces in low-temperature annealing district 1, thermophilic extension area 2 and the high-temperature denatured district 3 of controlled temperature respectively on described low-temperature annealing thermostatted, thermophilic extension thermostatted and the high-temperature denatured thermostatted; In low-temperature annealing district 1, thermophilic extension area 2 and high-temperature denatured district 3 have three required temperature of basic PC R reactions step respectively, above-mentioned three independently thermostatted above be placed with PCR micro-fluidic chip 6, in the PCR micro-fluidic chip, be distributed with the amplified reaction pipe; The amplified reaction pipe has PCR reaction fluid inlet 4, PCR reaction amplification back liquid outlet 9 and dna amplification reaction pipe; PCR reaction fluid inlet 4 is positioned at high-temperature denatured district 3 and is connected with miniature syringe pump, and the amplified reaction pipe passes high-temperature denatured district 3, thermophilic extension area 2 and low-temperature annealing district 1 successively, and the amplified reaction pipe is as the pcr amplification reaction district 5 in the PCR micro-fluidic chip; In the dna amplification reaction pipe of thermophilic extension area 2, be provided with electrochemical detector 8, the PCR reaction in the PCR micro-fluidic chip 6 can realize in each district in low-temperature annealing district 1, thermophilic extension area 2 and high-temperature denatured district 3; In the PCR micro-fluidic chip 6 high-temperature denatured district 3, thermophilic extension area 2 and the low-temperature annealing district 1 of each PCR reaction unwind with control, corresponding about extension and three accurate thermostatteds of annealed; It is inferior with loop cycle in the time of each humidity province with control PCR reaction solution that the pipeline of DNA cloning pipe is provided with the set flow velocity of length and pipeline inner fluid; The electrochemical detector of said thermophilic extension area 2 detects DNA electrochemistry hybridization indicator variable quantity in the PCR reaction amplification procedure, the amount of indirect reaction PCR reaction back amplified production, and the detection by quantitative of realization PCR reaction amplified material.
Described electrochemical detector comprises working electrode 10, counter electrode 11 and reference electrode 12, and working electrode 10, counter electrode 11 and reference electrode 12 guide line respectively are connected with testing circuit.
The making of the utility model device comprises following 3 steps:
(1)On firm banking, make at first again low-temperature annealing thermostatted, thermophilic extend thermostatted and high-temperature denatured thermostatted have three primitive reaction steps of pcr amplification temperature control (1, the low-temperature annealing district; 2, the thermophilic extension area; 3, high-temperature denatured district);
(2)Make the PCR micro-fluidic chip again, in the micro-fluidic chip microchannel length and rate of flow of fluid control the PCR reaction solution time of each humidity province and loop cycle number of times (5, the PCR expansion reaction zone in the micro-fluidic chip; 6, micro-fluidic chip); The thermostatted that each PCR conversion zone and accurate control are unwind, annealed and extends in the chip is corresponding up and down;
(3)The making of integrated electrochemical detector in the micro-fluidic chip, make at the thermophilic extension area microchannel place of micro-fluidic chip pcr amplification the integrated electrochemical detector (10, working electrode; 11, the counter electrode hole; 12, the reference electrode hole).
The temperature controlled thermostatted of three basic pcr amplification reaction steps adopts copper (or aluminium) matter metallic substance; The micro constant-temperature device is a rectangle; The PCR micro-fluidic chip is at three independently above the thermostatted, and is corresponding about each PCR temperature of reaction zone and the accurate thermostatted that control is unwind, annealed and extends in the PCR micro-fluidic chip.Metal thermostatted top is provided with electric heater, and the bottom is provided with scatterer, can carry out accurate temperature controlling to the temperature control working medium of thermostatted through modes such as PID.
Experimentation: earlier the quantitative PCR micro flow control chip device is assembled by above-mentioned steps; Open the constant-temperature temperature-control system of three thermostatteds again, regulate required experimental temperature (circulation sex change 95oC-annealing 60oC-extension 72oC), guarantee each PCR conversion zone homo(io)thermism in the PCR micro-fluidic chip; Utilize miniature syringe pump that PCR reaction solution (dna profiling, 4 * dNTPs and Taq-DNA polysaccharase, PCR damping fluid, BSA, DNA electrochemistry hybridization indicator etc.) is got into microchannel in ingress then, the PCR reaction solution can be controlled through microchannel length and rate of flow of fluid with loop cycle is inferior in the time of each humidity province; Utilize the integrated electrochemical detector to detect DNA electrochemistry hybridization indicator variable quantity in the PCR reaction amplification procedure at last, the amount of indirect reaction PCR reaction back amplified production, as accompanying drawing 3 (13, signal lines in the time of before the PCR reaction expansion; 14, PCR reaction expansion back signal lines), thereby the detection by quantitative of realization PCR reactant.
Making the quantitative PCR micro flow control chip device can carry out as follows: at first make temperature platform; Place the PCR micro-fluidic chip at temperature platform again, and it is inferior in the time and the loop cycle of each humidity province to utilize microchannel length and rate of flow of fluid to control the PCR reaction solution; Utilize the integrated electrochemical detector to detect DNA electrochemistry hybridization indicator variable quantity in the PCR reaction amplification procedure at last, the amount of indirect reaction PCR reaction back amplified production, and the detection by quantitative of realization PCR reaction amplified material.
Claims (3)
1. one kind
The quantitative PCR micro flow control chip deviceComprise independently thermostatted of firm banking (7), three; PCR micro-fluidic chip and electrochemical detector; It is characterized in that on firm banking (7), being manufactured with three independently thermostatteds; Three independently thermostatted is respectively the low-temperature annealing thermostatted, thermophilic extends thermostatted and high-temperature denatured thermostatted; Described low-temperature annealing thermostatted, thermophilic extend thermostatted and high-temperature denatured thermostatted is arranged side by side in regular turn; Described low-temperature annealing thermostatted, thermophilic extend three humidity provinces of the low-temperature annealing district (1) that is formed with accurate controlled temperature on thermostatted and the high-temperature denatured thermostatted respectively, thermophilic extension area and high-temperature denatured district (3), have three required temperature of basic PC R reactions step respectively at low-temperature annealing district (1), thermophilic extension area (2) and high-temperature denatured district (3), above-mentioned three independently thermostatted above be placed with PCR micro-fluidic chip (6); In the PCR micro-fluidic chip, be distributed with the amplified reaction pipe; The amplified reaction pipe has PCR reaction fluid inlet, PCR reaction amplification back liquid outlet (9) and dna amplification reaction pipe, and PCR reaction fluid inlet (4) is positioned at high-temperature denatured district and is connected with miniature syringe pump, and the amplified reaction pipe passes high-temperature denatured district (3), thermophilic extension area (2) and low-temperature annealing district (1) successively; The amplified reaction pipe is as the pcr amplification reaction district (5) in the PCR micro-fluidic chip, and the PCR reaction in the PCR micro-fluidic chip (6) can be accomplished in low-temperature annealing district (1), thermophilic extension area (2) and high-temperature denatured district (3); In the dna amplification reaction pipe of thermophilic extension area (2), be provided with the electrochemical detector (8) that the detection by quantitative of PCR reactant is used.
2. quantitative PCR micro flow control chip device according to claim 1, it is characterized in that described three independently thermostatted be the miniature rectangular thermostatted of processing by copper, aluminium or stainless material.
3. quantitative PCR micro flow control chip device according to claim 1 and 2 is characterized in that described electrochemical detector comprises working electrode (10), counter electrode (11) and reference electrode (12).
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| Application Number | Priority Date | Filing Date | Title |
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| CN 201220018703 CN202415561U (en) | 2012-01-16 | 2012-01-16 | Quantitative PCR (Polymerase Chain Reaction) microfluidic control chip device |
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| CN 201220018703 CN202415561U (en) | 2012-01-16 | 2012-01-16 | Quantitative PCR (Polymerase Chain Reaction) microfluidic control chip device |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102559488A (en) * | 2012-01-16 | 2012-07-11 | 福建医科大学 | Quantitative polymerase chain reaction (PCR) microfluidic chip integrated device for integrated electrochemical detection technology |
| CN102899245A (en) * | 2012-10-10 | 2013-01-30 | 凯晶生物科技(苏州)有限公司 | Micro-chamber static-state PCR (Polymerase Chain Reaction) and CE (Capillary Electrophoresis) function integrated micro-fluidic chip |
| CN104893954A (en) * | 2015-05-29 | 2015-09-09 | 上海交通大学 | Three-temperature-region channel embedded continuous flow laminated PCR (polymerase chain reaction) chip |
| CN107189938A (en) * | 2017-06-27 | 2017-09-22 | 中国科学院长春光学精密机械与物理研究所 | A kind of device quantitative for unknown nucleic acid concentration sample P CR |
| CN110872557A (en) * | 2018-08-30 | 2020-03-10 | 深圳市刚竹医疗科技有限公司 | PCR centrifugal microfluidic device and method thereof |
| CN115322888A (en) * | 2022-09-14 | 2022-11-11 | 翊新诊断技术(苏州)有限公司 | Microfluidic bag type circulating PCR chip and application thereof |
| WO2023005796A1 (en) * | 2021-07-28 | 2023-02-02 | 京东方科技集团股份有限公司 | Digital microfluidic apparatus and driving method therefor |
-
2012
- 2012-01-16 CN CN 201220018703 patent/CN202415561U/en not_active Expired - Lifetime
Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102559488A (en) * | 2012-01-16 | 2012-07-11 | 福建医科大学 | Quantitative polymerase chain reaction (PCR) microfluidic chip integrated device for integrated electrochemical detection technology |
| CN102899245A (en) * | 2012-10-10 | 2013-01-30 | 凯晶生物科技(苏州)有限公司 | Micro-chamber static-state PCR (Polymerase Chain Reaction) and CE (Capillary Electrophoresis) function integrated micro-fluidic chip |
| CN104893954A (en) * | 2015-05-29 | 2015-09-09 | 上海交通大学 | Three-temperature-region channel embedded continuous flow laminated PCR (polymerase chain reaction) chip |
| CN107189938A (en) * | 2017-06-27 | 2017-09-22 | 中国科学院长春光学精密机械与物理研究所 | A kind of device quantitative for unknown nucleic acid concentration sample P CR |
| CN107189938B (en) * | 2017-06-27 | 2020-09-08 | 中国科学院长春光学精密机械与物理研究所 | Device for PCR quantification of unknown nucleic acid concentration sample |
| CN110872557A (en) * | 2018-08-30 | 2020-03-10 | 深圳市刚竹医疗科技有限公司 | PCR centrifugal microfluidic device and method thereof |
| CN110872557B (en) * | 2018-08-30 | 2024-04-02 | 深圳市呈晖医疗科技有限公司 | PCR centrifugal microfluidic device and method thereof |
| WO2023005796A1 (en) * | 2021-07-28 | 2023-02-02 | 京东方科技集团股份有限公司 | Digital microfluidic apparatus and driving method therefor |
| CN115322888A (en) * | 2022-09-14 | 2022-11-11 | 翊新诊断技术(苏州)有限公司 | Microfluidic bag type circulating PCR chip and application thereof |
| CN115322888B (en) * | 2022-09-14 | 2024-01-19 | 翊新诊断技术(苏州)有限公司 | Microfluidic bag-type circulating PCR chip and application thereof |
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