CN110452802A

CN110452802A - It is complete to extract molecular diagnosis micro-fluidic chip and microfluidic system

Info

Publication number: CN110452802A
Application number: CN201910726228.1A
Authority: CN
Inventors: 汤明辉
Original assignee: Shenzhen City Of Gang Zhu Medical Science And Technology Co Ltd
Current assignee: Shenzhen Chenghui Medical Technology Co ltd
Priority date: 2019-08-07
Filing date: 2019-08-07
Publication date: 2019-11-15
Anticipated expiration: 2039-08-07
Also published as: CN110452802B

Abstract

Molecular diagnosis micro-fluidic chip and microfluidic system are extracted this application involves complete, the full molecular diagnosis micro-fluidic chip that extracts includes storage chamber, sample cavity, the first liquid feeding chamber, the first cushion chamber, at least one controlled liquid feeding chamber, the second liquid feeding chamber, distribution chamber, the first waste liquid chamber, cushion chamber, the second waste liquid chamber, at least one measurement chamber and an at least PCR amplification chamber；Controlled liquid feeding chamber is connected in the second waste liquid chamber and gas pipeline by gas pipeline at close-target rotation center and is equipped with phase transformation valve, and controlled liquid feeding chamber is connected to distribution chamber by charging line.The equivalent of sample cracking, nucleic acid purification, reagent is distributed, the nucleic acid of multi-chamber all processes such as amplification under specific primer constraint are all integrated in a centrifugal microfluidic control chip, and thus, it is possible to realize that the subsequent analysis such as the acquisition of fluorescence signal and analysis work, the diagnostic mode of single sample multi objective is realized, also realizes that more pathogen screenings in face of illness provide possibility for molecular diagnosis.

Description

It is complete to extract molecular diagnosis micro-fluidic chip and microfluidic system

Technical field

This application involves centrifugal microfluidic control fields, extract molecular diagnosis micro-fluidic chip and micro-fluidic system more particularly to complete System.

Background technique

Micro-fluidic (Microfluidics) refers to the handling liquids on submillimeter scale.It is by biological and chemical field institute The basic operation unit being related to even collects the function in entire laboratory, including sampling, dilution, reaction, separation, detection etc. On Cheng Yi small chip, therefore also known as chip lab (Lab-on-a-Chip).This chip is usually by various liquid storage tanks It is formed with microchannel network interconnected, can largely shorten the sample process time, and control liquid flowing by accurate, Realize the maximum utilization efficiency of reagent consumptive material.It is micro-fluidic for biomedical research, pharmaceutical synthesis screening, environmental monitoring and protection, The application of the various fields such as health quarantine, judicial expertise, the detection of biological reagent provides extremely wide prospect.Particularly, The micro-fluidic demand that can meet care diagnostic (Point-of-care testing, POCT) miniaturization instrument well, so by It is widely applied in POCT.In industrialization, micro-fluidic to be generally divided into following several big types: pressure (air pressure or hydraulic) is driven Dynamic formula is micro-fluidic, centrifugal microfluidic control, drop is micro-fluidic, digitlization is micro-fluidic, papery is micro-fluidic etc..

Microfluidic system refers to the device of the handling liquids on submillimeter scale (general several microns to several hundred microns).From The heart is micro-fluidic to be under the jurisdiction of a micro-fluidic branch, refer in particular to by rotation centrifugal microfluidic control chip come using centrifugal force in submillimeter The flowing of handling liquids on scale.Basic operation unit involved in biological and chemical field is integrated in a small-sized dish-style by it (disc-shaped) chip on.Other than micro-fluidic specific advantage, since centrifugal microfluidic control only needs a motor Power required for liquid manipulation is provided, so whole equipment more concision and compact.And on disc-type chip it is ubiquitous from Psychological field can make liquid driven more effective, it is ensured that without residual liquid in pipeline, and can effectively realize based on density contrast Different sample separation, also can allow parallel processing more simple.Therefore, centrifugal microfluidic control is also more and more applied and is examined immediately In disconnected.

The molecular diagnosis of based on PCR amplification is that specific amplification target gene is mediated (to lose to detect endogenous by primer Pass or variation) or exogenous (pathogen) target gene presence or absence, and then to the diagnosing and treating of disease provide information and Decision-making foundation.Its main application scenarios has the diagnosis of infectious disease, blood sieve, Tumor mutations site primer, the diagnosis of hereditary disease, production Preceding diagnosis, tissue typing etc..The molecular diagnosis of based on PCR amplification generally comprises following steps: sample cracking, nucleic acid purification, core Acid expands under specific primer constraint, the acquisition and analysis of fluorescence signal.

But based on PCR amplification molecular diagnosis system in, due to having Aerosol Pollution when PCR amplification, also for The cross contamination between sample is avoided, to set up a subregion laboratory under normal circumstances.This laboratory will realize sample Processing, nucleic acid extraction, the division operation of PCR amplification, and must have good ventilating system, laboratory build it is at high cost, it is past Just there are the financial resources built toward only larger medical mechanism.On the other hand, laboratory operation personnel will take appointment with certificate, and also greatly increase Cost of labor.At the same time, excessive artificial intervention certainly will also bring along artificial operation error.This is all greatly increased The technology of the molecular diagnosis of based on PCR uses threshold.Also, current molecular diagnostic laboratories mode is concentrating experimental site Multisample and the operation of more detection projects are completed, process quality control requires high.Although molecular diagnostic techniques advantage is it is obvious that still Due to its complex steps, process is time-consuming, and professional is needed to operate, and clinical molecular diagnosis laboratory to build cost general It is higher, so molecular diagnosis is also expensive.And current molecular diagnostic laboratories mode, generally multisample single index are examined Survey mode, Testing index is limited, cannot achieve the screening of single sample multi objective pathogen infection.

Applicant has been proposed exempting from thus the related micro fluidic chip technology of nucleic acid purification, but there are many more detections Mesh inevitably carries out nucleic acid purification.

Summary of the invention

Based on this, it is necessary to provide a kind of extraction molecular diagnosis micro-fluidic chip and microfluidic system entirely, mainly solve such as What extracts the problem of molecular diagnosis is integrated in a centrifugal microfluidic control chip for complete, by sample cracking, nucleic acid purification, nucleic acid in spy Determine all processes such as the lower amplification of primer constraint to be all integrated in a centrifugal microfluidic control chip, and is achieved in adopting for fluorescence signal Collection and analysis etc..

A kind of full extraction molecular diagnosis micro-fluidic chip, with target rotation center, the full extraction molecular diagnosis is micro- Fluidic chip include storage chamber, sample cavity, the first liquid feeding chamber, the first cushion chamber, at least one controlled liquid feeding chamber, the second liquid feeding chamber, Distribute chamber, the first waste liquid chamber, cushion chamber, the second waste liquid chamber, at least one measurement chamber and an at least PCR amplification chamber；

By the sequence arrangement at a distance from the target rotation center from small to large are as follows: storage chamber, sample cavity, the first buffering Chamber, controlled liquid feeding chamber, distribution chamber, the first waste liquid chamber, measurement chamber, PCR amplification chamber；

First liquid feeding chamber is greater than storage chamber at a distance from target rotation center and less than the at a distance from target rotation center One cushion chamber is at a distance from target rotation center；

It is at a distance from target rotation center and small that second liquid feeding chamber is greater than the first cushion chamber at a distance from target rotation center In distribution chamber at a distance from target rotation center；

It is at a distance from target rotation center and small that cushion chamber is more than or equal to the first waste liquid chamber at a distance from target rotation center In measurement chamber at a distance from target rotation center；

Second waste liquid chamber is greater than cushion chamber at a distance from target rotation center at a distance from target rotation center；

Storage chamber is equipped with the first well at close-target rotation center, and storage chamber is connected to sample by the first capillary valves Chamber；

Sample cavity is equipped with the second well at close-target rotation center, and sample cavity passes through the first siphon piping connection first Cushion chamber；

First liquid feeding chamber is equipped with the first liquid filling hole at close-target rotation center, and the first liquid feeding chamber passes through the second siphon piping It is connected to the first cushion chamber；

First cushion chamber is equipped with the first cushion hole at close-target rotation center, and the first cushion chamber passes through third siphon piping Connection distribution chamber；

Second liquid feeding chamber is equipped with the second liquid filling hole at close-target rotation center, and the second liquid feeding chamber passes through elution pipeline connection Distribute chamber；

Controlled liquid feeding chamber is connected in the second waste liquid chamber and gas pipeline by gas pipeline at close-target rotation center and is set There is phase transformation valve, controlled liquid feeding chamber is connected to distribution chamber by charging line；

The full molecular diagnosis micro-fluidic chip that extracts is in the inlet or distribution chamber of distributing in chamber or distribute chamber Entrance before be equipped with filter structure；

Distribute chamber and the first waste liquid chamber is connected to by waste solution channel far from one end at target rotation center, the other end passes through Collection conduit is connected to the second cushion chamber, and waste solution channel and collection conduit are located at the two sides of distribution chamber；

First waste liquid chamber is equipped with waste liquid venthole at close-target rotation center；

Second cushion chamber is connected to sample by the 4th siphon piping and distributes pipeline；

Sample distribution pipeline sequential communication respectively measures chamber and end is connected to the second waste liquid chamber；Measure chamber and PCR amplification chamber Quantity is identical and is correspondingly arranged, and each measurement chamber is connected to a PCR amplification chamber；

Second waste liquid chamber is equipped with venthole at close-target rotation center.

Above-mentioned full extraction molecular diagnosis micro-fluidic chip, by the equivalent distribution of sample cracking, nucleic acid purification, reagent, multi-cavity The nucleic acid of room all processes such as amplification under specific primer constraint are all integrated in a centrifugal microfluidic control chip, and thus, it is possible to It realizes that the subsequent analysis such as acquisition and the analysis of fluorescence signal work, realizes the diagnostic mode of single sample multi objective, be also molecule Diagnosis realizes that more pathogen screenings in face of illness provide possibility；Also, entire reaction process is in closed micro-fluidic core In piece, a possibility that reducing the burden and pollution of operator, but also entire molecular diagnostic procedure is no longer dependent on molecule Diagnostic test room, is also no longer dependent on professional operator, realizes the demand quickly detected whenever and wherever possible, is medical inspection Huge help is brought with control and prevention of disease.

The full extraction molecular diagnosis micro-fluidic chip includes two controlled liquid feeding chambers in one of the embodiments, respectively For the first controlled liquid feeding chamber and the second controlled liquid feeding chamber；

By the sequence arrangement at a distance from the target rotation center from small to large are as follows: storage chamber, sample cavity, the first buffering Chamber, the first controlled liquid feeding chamber, the second controlled liquid feeding chamber, distribution chamber, the first waste liquid chamber, measurement chamber, PCR amplification chamber；

First controlled liquid feeding chamber is connected to the second waste liquid chamber and first by first gas pipeline at close-target rotation center The first paraffin valve is equipped in gas pipeline, the first controlled liquid feeding chamber passes through the first charging line connection distribution chamber；

Second controlled liquid feeding chamber is connected to the second waste liquid chamber and second by second gas pipeline at close-target rotation center The second paraffin valve is equipped in gas pipeline, the second controlled liquid feeding chamber passes through the second charging line connection distribution chamber.

In one of the embodiments, the full extraction molecular diagnosis micro-fluidic chip further include convergence chamber, by with it is described The sequence arrangement of the distance of target rotation center from small to large are as follows: controlled liquid feeding chamber, convergence chamber, distribution chamber；

It is at a distance from target rotation center and small that second liquid feeding chamber is greater than the first cushion chamber at a distance from target rotation center In convergence chamber at a distance from target rotation center；

First cushion chamber passes through third siphon piping connection convergence chamber；

Second liquid feeding chamber passes through elution pipeline connection convergence chamber；

Controlled liquid feeding chamber is connected to convergence chamber by charging line；

It converges chamber and passes through the second capillary valves connection distribution chamber.

The first cushion chamber is connected to collection conduits, the second liquid feeding chamber by third siphon piping in one of the embodiments, Collection conduits are connected to by elution pipeline, controlled liquid feeding chamber is connected to collection conduits, collection conduits connection convergence by charging line Chamber.

It converges in chamber in one of the embodiments, and is equipped with filter membrane as the filter structure；Further, the filter Film is the multilayer pellosil being stacked.

By the sequence arrangement at a distance from the target rotation center from small to large are as follows: storage chamber, sample cavity, the first buffering Chamber, the first controlled liquid feeding chamber, the second controlled liquid feeding chamber, convergence chamber, distribution chamber, the first waste liquid chamber, measurement chamber, PCR amplification chamber；

First controlled liquid feeding chamber is connected to the second waste liquid chamber and first by first gas pipeline at close-target rotation center The first paraffin valve is equipped in gas pipeline, the first controlled liquid feeding chamber passes through the first charging line connection convergence chamber；

Second controlled liquid feeding chamber is connected to the second waste liquid chamber and second by second gas pipeline at close-target rotation center The second paraffin valve is equipped in gas pipeline, the second controlled liquid feeding chamber passes through the second charging line connection convergence chamber；

The first cushion chamber is connected to collection conduits, the second liquid feeding chamber by third siphon piping in one of the embodiments, Collection conduits are connected to by elution pipeline, the first controlled liquid feeding chamber is connected to collection conduits by the first charging line, and second is controlled Liquid feeding chamber is connected to collection conduits by the second charging line；Collection conduits connection convergence chamber.

The full extraction molecular diagnosis micro-fluidic chip further includes reagent conveyance conduit in one of the embodiments, is surveyed Amount chamber is identical as the quantity of reagent conveyance conduit and is correspondingly arranged, and each measurement chamber is connected to by corresponding reagent conveyance conduit Corresponding PCR amplification chamber；Alternatively, the end of sample distribution pipeline is connected to the second waste liquid chamber by Drainage pipe.

The full extraction molecular diagnosis micro-fluidic chip has ontology in one of the embodiments, and the ontology opens up There are storage chamber, the first capillary valves, sample cavity, Drainage pipe, the first siphon piping, the second siphon piping, the first liquid feeding chamber, second Liquid feeding chamber, the first cushion chamber, third siphon piping, collection conduits, the first controlled liquid feeding chamber, the second controlled liquid feeding chamber, the first gas Body pipeline, second gas pipeline, the first charging line, the second charging line, elution pipeline, convergence chamber, the second capillary valves, distribution Chamber, waste solution channel, collection conduit, the second cushion chamber, the 4th siphon piping, the first waste liquid chamber, sample distribution pipeline, each measurement Some or all of in chamber, each PCR amplification chamber, the second waste liquid chamber and reagent conveyance conduit；First well, the second well, First liquid filling hole, the first cushion hole, the second liquid filling hole, waste liquid venthole and venthole are each passed through the ontology and connection is external Environment；And/or magnetic part and/or abrasive article are equipped in sample cavity；And/or paraffin is preset in PCR amplification chamber 135.

The first well, the second well, the first liquid filling hole, the first cushion hole, second add in one of the embodiments, Fluid apertures, waste liquid venthole and venthole are each passed through the same side of the ontology and connection external environment.

A kind of microfluidic system comprising extract molecular diagnosis micro-fluidic chip described in any of the above-described entirely.

Above-mentioned microfluidic system distributes the equivalent of sample cracking, nucleic acid purification, reagent, the nucleic acid of multi-chamber is specific The all processes such as the lower amplification of primer constraint are all integrated in a centrifugal microfluidic control chip, and thus, it is possible to realize fluorescence signal The subsequent analysis such as acquisition and analysis work, and realize the diagnostic mode of single sample multi objective, also realize for molecular diagnosis in face of disease More pathogen screenings of disease provide possibility；Also, entire reaction process is in closed micro-fluidic chip, reduces operation A possibility that burden and pollution of personnel, but also entire molecular diagnostic procedure is no longer dependent on molecular diagnostic laboratories, also not Professional operator is depended on again, realizes the demand quickly detected whenever and wherever possible, is brought for medical inspection and control and prevention of disease Huge help.

Detailed description of the invention

Fig. 1 is the schematic diagram of internal structure in a direction of one embodiment of the application.

Fig. 2 is the schematic diagram of another direction of embodiment illustrated in fig. 1.

Fig. 3 is the schematic diagram of another direction of embodiment illustrated in fig. 1.

Fig. 4 is the schematic diagram of another direction of embodiment illustrated in fig. 1.

Fig. 5 is the schematic diagram of another direction of embodiment illustrated in fig. 1.

Specific embodiment

In order to make the above objects, features, and advantages of the present application more apparent, with reference to the accompanying drawing to the application Specific embodiment be described in detail.Many details are explained in the following description in order to fully understand this Shen Please.But the application can be implemented with being much different from other way described herein, those skilled in the art can be not Similar improvement is done in the case where violating the application intension, therefore the application is not limited by the specific embodiments disclosed below.

It should be noted that it can be directly another when element is referred to as " being fixed on " or " being set to " another element On one element or there may also be elements placed in the middle.When an element is considered as " connection " another element, it can be with It is directly to another element or may be simultaneously present centering elements.Term as used herein " vertically ", " level ", "left", "right" and similar statement for illustrative purposes only, be not meant to be the only embodiment.

Unless otherwise defined, all technical and scientific terms used herein and the technical field for belonging to the application The normally understood meaning of technical staff is identical.The term used in the description of the present application is intended merely to description tool herein The purpose of the embodiment of body, it is not intended that in limitation the application.Term " and or " used herein includes one or more Any and all combinations of relevant listed item.

It is a kind of complete to extract molecular diagnosis micro-fluidic chip in the application one embodiment, with target rotation center, The full extraction molecular diagnosis micro-fluidic chip include storage chamber, sample cavity, the first liquid feeding chamber, the first cushion chamber, at least one by Control liquid feeding chamber, the second liquid feeding chamber, distribution chamber, the first waste liquid chamber, cushion chamber, the second waste liquid chamber, at least one measurement chamber and at least One PCR amplification chamber；By the sequence arrangement at a distance from the target rotation center from small to large are as follows: storage chamber, sample cavity, first Cushion chamber, controlled liquid feeding chamber, distribution chamber, the first waste liquid chamber, measurement chamber, PCR amplification chamber；In first liquid feeding chamber and target rotation The distance of the heart be greater than storage chamber at a distance from target rotation center and less than the first cushion chamber at a distance from target rotation center；The Two liquid feeding chambers are greater than the first cushion chamber at a distance from target rotation center at a distance from target rotation center and are less than distribution chamber At a distance from target rotation center；Cushion chamber is more than or equal in the first waste liquid chamber and target rotation at a distance from target rotation center The distance of the heart and be less than measurement chamber at a distance from target rotation center；Second waste liquid chamber is greater than slow at a distance from target rotation center Chamber is rushed at a distance from target rotation center；Storage chamber is equipped with the first well at the close-target rotation center, and storage chamber passes through the One capillary valves are connected to sample cavity；Sample cavity is equipped with the second well at close-target rotation center, and sample cavity passes through the first siphon Pipeline is connected to the first cushion chamber；First liquid feeding chamber is equipped with the first liquid filling hole at close-target rotation center, and the first liquid feeding chamber passes through Second siphon piping is connected to the first cushion chamber；First cushion chamber is equipped with the first cushion hole at close-target rotation center, and first is slow It rushes chamber and passes through third siphon piping connection distribution chamber；Second liquid feeding chamber is equipped with the second liquid filling hole at close-target rotation center, Second liquid feeding chamber passes through elution pipeline connection distribution chamber；Controlled liquid feeding chamber is connected at close-target rotation center by gas pipeline Phase transformation valve is equipped in logical second waste liquid chamber and gas pipeline, controlled liquid feeding chamber is connected to distribution chamber by charging line；It is described complete Molecular diagnosis micro-fluidic chip is extracted to be equipped in distribution chamber or before distributing the inlet of chamber or the entrance of distribution chamber Filter structure；Distribute chamber and the first waste liquid chamber is connected to by waste solution channel far from one end at target rotation center, the other end passes through Collection conduit is connected to the second cushion chamber, and waste solution channel and collection conduit are located at the two sides of distribution chamber；First waste liquid chamber Waste liquid venthole is equipped at close-target rotation center；Second cushion chamber is connected to sample by the 4th siphon piping and distributes pipeline； Sample distribution pipeline sequential communication respectively measures chamber and end is connected to the second waste liquid chamber；It is identical as the quantity of PCR amplification chamber to measure chamber And be correspondingly arranged, each measurement chamber is connected to a PCR amplification chamber；Second waste liquid chamber is equipped with venthole at close-target rotation center. Above-mentioned full extraction molecular diagnosis micro-fluidic chip, by the equivalent distribution of sample cracking, nucleic acid purification, reagent, the nucleic acid of multi-chamber The all processes such as amplification are all integrated in a centrifugal microfluidic control chip under specific primer constraint, and thus, it is possible to realize fluorescence The subsequent analysis such as the acquisition and analysis of signal work, and realize the diagnostic mode of single sample multi objective, are also molecular diagnosis realization More pathogen screenings in face of illness provide possibility；Also, entire reaction process is in closed micro-fluidic chip, is reduced A possibility that burden and pollution of operator, but also entire molecular diagnostic procedure is no longer dependent on molecular diagnostic assays Room is also no longer dependent on professional operator, realizes the demand quickly detected whenever and wherever possible, is that medical inspection and disease are anti- Control brings huge help.

A kind of full extraction molecular diagnosis micro-fluidic chip in one of the embodiments, comprising the portion of following embodiment Separation structure or entire infrastructure；That is, extracting molecular diagnosis micro-fluidic chip entirely includes some technical characteristics below or whole technologies Feature.In one of the embodiments, the full extraction molecular diagnosis micro-fluidic chip include storage chamber, sample cavity, first plus Sap cavity, the first cushion chamber, at least one controlled liquid feeding chamber, the second liquid feeding chamber, distribution chamber, the first waste liquid chamber, cushion chamber, second are given up Sap cavity, at least one measurement chamber and an at least PCR amplification chamber.The target rotation center is used in one of the embodiments, As rotation center when centrifugation.Target rotation center can be entity, be also possible to diastema；Target rotation center can mention entirely It takes inside molecular diagnosis micro-fluidic chip, can also extract outside molecular diagnosis micro-fluidic chip entirely, but usually illusory in complete It extracts outside molecular diagnosis micro-fluidic chip, that is, target rotation center is the opposite object of reference an of outside.One wherein In embodiment, the full molecular diagnosis micro-fluidic chip that extracts is for being set in microfluidic system, and target rotation center is miniflow The centrifugation center of control system.Further, the target rotation center is located at the full extraction point in one of the embodiments, Inside son diagnosis micro-fluidic chip or the target rotation center is located at outside the full extraction molecular diagnosis micro-fluidic chip Portion.In this way, it is only necessary to which microfluidic system or its ontology are in constant centrifugal rotational speed and can realize the sequence load of reagent.

It is described at a distance from the target rotation center in each embodiment, including the minimum with the target rotation center Perhaps center center at a distance from the target rotation center includes geometric center or mass center etc. to distance.That is, right In storage chamber at a distance from the target rotation center, minimum range or storage including storage chamber Yu the target rotation center The center of chamber is deposited at a distance from the target rotation center；It is described in one of the embodiments, to be rotated with the target The distance at center is the minimum range with the target rotation center.It is described in one of the embodiments, to be revolved with the target The distance for turning center is center position at a distance from the target rotation center.Each embodiment of the application does not make special limit to this System need to only can be realized centrifugal microfluidic control；Those skilled in the art can select according to actual needs it is described with it is described The distance of target rotation center is to realize the full extraction molecular diagnosis micro-fluidic chip.It is understood that except through each A hole is communicated to except external environment, each chamber be actually for external environment it is closed, each hole include first Well, the second well, the first liquid filling hole, the first cushion hole, the second liquid filling hole, waste liquid venthole and venthole etc. consider To the effect of centrifugal force, in each embodiment, the position in each hole is generally located near at target rotation center and avoids each pipeline Communication port；Each pipeline includes the first capillary valves, Drainage pipe, the first siphon piping, the second siphon piping, third siphon pipe Road, collection conduits, first gas pipeline, second gas pipeline, the first charging line, the second charging line, elution pipeline, second Capillary valves, waste solution channel, collection conduit, the 4th siphon piping, sample distribute pipeline and reagent conveyance conduit etc.；Each chamber includes Storage chamber, sample cavity, the first liquid feeding chamber, the second liquid feeding chamber, the first cushion chamber, the first controlled liquid feeding chamber, the second controlled liquid feeding chamber, It converges chamber, distribution chamber, the second cushion chamber, the first waste liquid chamber, respectively measure chamber, the portion in each PCR amplification chamber and the second waste liquid chamber Divide or whole.Alternatively, each chamber includes storage chamber, the first capillary valves, sample cavity, Drainage pipe, the first siphon piping, the second rainbow Suction pipe road, the first liquid feeding chamber, the second liquid feeding chamber, the first cushion chamber, third siphon piping, collection conduits, the first controlled liquid feeding chamber, Second controlled liquid feeding chamber, first gas pipeline, second gas pipeline, the first charging line, the second charging line, elution pipeline, Converge chamber, the second capillary valves, distribution chamber, waste solution channel, collection conduit, the second cushion chamber, the 4th siphon piping, the first waste liquid Chamber, sample distribution pipeline respectively measure chamber, each PCR amplification chamber, some or all of in the second waste liquid chamber and reagent conveyance conduit. Pipeline is substantially exactly long and narrow small chamber.Further, distribution chamber is rectangle, ellipse in one of the embodiments, Or strip, distribute the length that chamber is greater than direction of rotation in the length of centrifugal direction in one of the embodiments, wherein In one embodiment, distribution chamber is longer than the design of convergence chamber, and the presence for distributing chamber is to increase Coriolis force effect In the time of liquid, to make liquid more reliable when doing channel conversion.

Further, in one of the embodiments, storage chamber, sample cavity, each cushion chamber, each liquid feeding chamber, it is each it is controlled plus Sap cavity and/or each measurement chamber are in it far from all having collapsed shape at target rotation center；In one of the embodiments, in Triangle, circle, shuttle shape or ellipse etc..Such design advantageously allows liquid and is exported from collapsed shape to realize and add Carry or export complete effect.Further, in one of the embodiments, each chamber in it far from the target rotation center Place is connected to corresponding chamber by associated conduit, and corresponding chamber is connected to associated conduit in it at close-target rotation center, In this way, being conducive to the output and input of liquid.

The full extraction molecular diagnosis micro-fluidic chip is equipped with filtering knot in distribution chamber in one of the embodiments, Structure；Or the full extraction molecular diagnosis micro-fluidic chip is equipped with filter structure in the inlet of distribution chamber；Or the full extraction Molecular diagnosis micro-fluidic chip is equipped with filter structure before the entrance of distribution chamber；Further, in one of the embodiments, The filter structure includes filter membrane, filter core or filter etc..Filter structure is set in distribution chamber or filter structure is set It is placed in front of distribution chamber, distributes after enabling distribution chamber to play the role of a filtering.

Further, the phase transformation valve is used to undergo phase transition by temperature control to be connected in one of the embodiments, The gas pipeline.In one of the embodiments, the full extraction molecular diagnosis micro-fluidic chip include at least two it is controlled plus Sap cavity, and the different setting of phase transition temperature of the phase transformation valve of different gas pipeline prepare phase using the material of different phase transition temperature The phase transformation valve of different gas pipeline, so that the liquid of each controlled liquid feeding chamber is added in an orderly manner in distribution chamber or convergence chamber.In In one embodiment, the full extraction molecular diagnosis micro-fluidic chip includes two controlled liquid feeding chambers, and respectively first is controlled Liquid feeding chamber and the second controlled liquid feeding chamber；By the sequence arrangement at a distance from the target rotation center from small to large are as follows: storage chamber, Sample cavity, the first cushion chamber, the first controlled liquid feeding chamber, the second controlled liquid feeding chamber, distribution chamber, the first waste liquid chamber, measurement chamber, PCR amplification chamber；First controlled liquid feeding chamber is connected to the second waste liquid chamber and the by first gas pipeline at the close-target rotation center The first paraffin valve is equipped in one gas pipeline, the first controlled liquid feeding chamber passes through the first charging line connection distribution chamber；Second by Control liquid feeding chamber is connected in the second waste liquid chamber and second gas pipeline by second gas pipeline at close-target rotation center and is equipped with Second paraffin valve, the second controlled liquid feeding chamber pass through the second charging line connection distribution chamber.That is, in one of the embodiments, A kind of full extraction molecular diagnosis micro-fluidic chip, with target rotation center, the full extraction molecular diagnosis micro-fluidic chip Add including storage chamber, sample cavity, the first liquid feeding chamber, the first cushion chamber, the first controlled liquid feeding chamber, the second controlled liquid feeding chamber, second Sap cavity, distribution chamber, the first waste liquid chamber, cushion chamber, the second waste liquid chamber, at least one measurement chamber and an at least PCR amplification chamber；By with The sequence arrangement of the distance of the target rotation center from small to large are as follows: storage chamber, sample cavity, the first cushion chamber, first controlled Liquid feeding chamber, the second controlled liquid feeding chamber, distribution chamber, the first waste liquid chamber, measurement chamber, PCR amplification chamber；First liquid feeding chamber and target are revolved The distance that turns center be greater than storage chamber with target rotation center at a distance from and less than the first cushion chamber and target rotation center away from From；Second liquid feeding chamber is greater than the first cushion chamber at a distance from target rotation center at a distance from target rotation center and is less than distribution Chamber is at a distance from target rotation center；Cushion chamber is more than or equal to the first waste liquid chamber at a distance from target rotation center and target is revolved Turn the distance at center and is less than measurement chamber at a distance from target rotation center；Second waste liquid chamber is big at a distance from target rotation center In cushion chamber at a distance from target rotation center；Storage chamber is equipped with the first well at close-target rotation center, and storage chamber is logical Cross the first capillary valves connection sample cavity；Sample cavity is equipped with the second well at close-target rotation center, and sample cavity passes through first Siphon piping is connected to the first cushion chamber；First liquid feeding chamber is equipped with the first liquid filling hole, the first liquid feeding chamber at close-target rotation center The first cushion chamber is connected to by the second siphon piping；First cushion chamber is equipped with the first cushion hole at close-target rotation center, the One cushion chamber passes through third siphon piping connection distribution chamber；Second liquid feeding chamber is equipped with the second liquid feeding at close-target rotation center Hole, the second liquid feeding chamber pass through elution pipeline connection distribution chamber；First controlled liquid feeding chamber is at the close-target rotation center by the One gas pipeline, which is connected in the second waste liquid chamber and first gas pipeline, is equipped with the first paraffin valve, and the first controlled liquid feeding chamber passes through first Charging line connection distribution chamber；Second controlled liquid feeding chamber passes through second gas pipeline connection second at close-target rotation center The second paraffin valve is equipped in waste liquid chamber and second gas pipeline, the second controlled liquid feeding chamber passes through the second charging line connection distribution chamber Room；The full molecular diagnosis micro-fluidic chip that extracts is in the entrance for distributing in the chamber or inlet of distribution chamber or distribution chamber It is preceding to be equipped with filter structure；Distribute chamber and the first waste liquid chamber is connected to by waste solution channel far from one end at target rotation center, separately One end is connected to the second cushion chamber by collection conduit, and waste solution channel and collection conduit are located at the two sides of distribution chamber；The One waste liquid chamber is equipped with waste liquid venthole at close-target rotation center；Second cushion chamber is connected to sample point by the 4th siphon piping Send out pipeline；Sample distribution pipeline sequential communication respectively measures chamber and end is connected to the second waste liquid chamber；Measure the number of chamber and PCR amplification chamber It measures identical and is correspondingly arranged, each measurement chamber is connected to a PCR amplification chamber；Second waste liquid chamber is equipped at close-target rotation center Stomata.Remaining embodiment and so on.Further, the full extraction molecular diagnosis micro-fluidic chip include it is greater number of by Liquid feeding chamber is controlled, in this way, being advantageously implemented the orderly controlled progress of multiple liquid feeding.Phase transformation valve and design, relative to common passive Valve, hydrophilic and hydrophobic and surface tension of the phase transformation valve independent of reagent have more universality, and repeatability and reliability also more can It is guaranteed；In this way, melting the phase-change material of different melting points sequentially by temperature control to realize the liquid examination of controlled liquid feeding chamber Agent is distributed in certain sequence in chamber or convergence chamber, it is only necessary to which the suitable of reagent can be realized by being in constant centrifugal rotational speed Sequence load；And it need to only simply change the sequence load that can realize the reagent in any controlled liquid feeding chamber.Relative to common Passive valve such as capillary valves, siphon valve, drain valve etc., hydrophilic and hydrophobic and surface tension of the phase transformation valve independent of reagent, more There is universality, repeatability and reliability also can more be guaranteed, and phase transformation valve uses phase-change material in one of the embodiments, It realizes, is realized in one of the embodiments, using phase-change material closing load pipeline or air flue or outlet pipe.Phase-change material Including but not limited to solid paraffin, synthetic wax, crystalline wax, native paraffin etc. and various thermoplastic materials such as polycarbonate (PC), PMMA, COC etc. and various room temperature are solid, the material etc. melted after appropriate heating.Phase-change material is solid in normal temperature state State, melting after heating is liquid, therefore plays the role of valve.Also it is important to note that the implementation of phase transformation valve It is varied, it common are phase-change material and directly block gas pipeline, heating phase transformation valve is controlled after melting its phase-change material to be added The venthole that sap cavity just passes through the second waste liquid chamber is connected with the atmospheric pressure of external environment, is just able to achieve the liquid that controlled liquid feeding is intracavitary Body is discharged into distribution chamber or convergence chamber.Such design melts the phase-change material of different melting points by change temperature, Realize that liquid breaks through different phase transformation valves in different time, to realize the sequence load of liquid reagent.As long as therefore passing through Temperature control melts the phase-change materials of different melting points to realize application that the liquid reagent of controlled liquid feeding chamber loads in certain sequence Mode should be understood as the protection scope for belonging to each embodiment of the application, and be not limited to the implementation of specific phase transformation valve In.

In one of the embodiments, the full extraction molecular diagnosis micro-fluidic chip further include convergence chamber, by with it is described The sequence arrangement of the distance of target rotation center from small to large are as follows: controlled liquid feeding chamber, convergence chamber, distribution chamber；Second liquid feeding chamber It is rotated with being greater than the first cushion chamber at a distance from target rotation center at a distance from target rotation center and being less than convergence chamber with target The distance at center；First cushion chamber passes through third siphon piping connection convergence chamber；Second liquid feeding chamber is converged by elution pipeline connection Poly- chamber；Controlled liquid feeding chamber is connected to convergence chamber by charging line；It converges chamber and passes through the second capillary valves connection distribution chamber.That is, each Liquid in chamber is entered in distribution chamber by converging chamber.It converges in one of the embodiments, and is equipped with filter membrane work in chamber For the filter structure；Further, the filter membrane is the multilayer pellosil being stacked in one of the embodiments,.

The first cushion chamber is connected to collection conduits, the second liquid feeding chamber by third siphon piping in one of the embodiments, Collection conduits are connected to by elution pipeline, controlled liquid feeding chamber is connected to collection conduits, collection conduits connection convergence by charging line Chamber.It is connected to convergence chamber again after the pipeline connection collection conduits of i.e. each chamber.The full extraction molecule in one of the embodiments, Diagnosing micro-fluidic chip includes two controlled liquid feeding chambers, the respectively first controlled liquid feeding chamber and the second controlled liquid feeding chamber；By with it is described The sequence arrangement of the distance of target rotation center from small to large are as follows: storage chamber, sample cavity, the first cushion chamber, the first controlled liquid feeding Chamber, the second controlled liquid feeding chamber, convergence chamber, distribution chamber, the first waste liquid chamber, measurement chamber, PCR amplification chamber；First controlled liquid feeding chamber It is connected in the second waste liquid chamber and first gas pipeline at close-target rotation center by first gas pipeline and is equipped with the first paraffin Valve, the first controlled liquid feeding chamber pass through the first charging line connection convergence chamber；Second controlled liquid feeding chamber is at close-target rotation center It is connected in the second waste liquid chamber and second gas pipeline by second gas pipeline and is equipped with the second paraffin valve, the second controlled liquid feeding chamber is logical Cross the second charging line connection convergence chamber；The first cushion chamber is converged by the connection of third siphon piping in one of the embodiments, Collect pipeline, the second liquid feeding chamber is connected to collection conduits by elution pipeline, and the first controlled liquid feeding chamber is connected to by the first charging line Collection conduits, the second controlled liquid feeding chamber are connected to collection conduits by the second charging line；Collection conduits connection convergence chamber.

The full extraction molecular diagnosis micro-fluidic chip further includes reagent conveyance conduit in one of the embodiments, is surveyed Amount chamber is identical as the quantity of reagent conveyance conduit and is correspondingly arranged, and each measurement chamber is connected to by corresponding reagent conveyance conduit Corresponding PCR amplification chamber；The end of sample distribution pipeline is useless by Drainage pipe connection second in one of the embodiments, Sap cavity.Further, the minimum range of Drainage pipe and target rotation center is more than or equal to sample in one of the embodiments, Distribute the maximum distance of pipeline and target rotation center.Further, the number of chamber, reagent conveyance conduit and PCR amplification chamber is measured Amount is 8.The application proposes a kind of full extraction molecular diagnosis micro-fluidic chip, belongs to a kind of centrifugal microfluidic control chip, cooperates Full-automatic foranalysis of nucleic acids instrument realizes the full full-automation for extracting molecular diagnosis project.In this micro-fluidic chip, sample The equivalent distribution of cracking, nucleic acid purification and reagent, the PCR amplification of multi-chamber are all able to sequence and realize.At entire reaction process In closed micro-fluidic chip, a possibility that reducing the burden and pollution of operator, but also entire molecular diagnosis mistake Journey is no longer dependent on molecular diagnostic laboratories, is also no longer dependent on professional operator, realizes whenever and wherever possible quickly detection Demand, bring huge help for medical inspection and control and prevention of disease.In the embodiment, this extracts the micro-fluidic core of molecular diagnosis entirely Piece has 8 PCR amplification chambers, and the corresponding foranalysis of nucleic acids instrument of each amplification chamber has 5 kinds of fluorescence channels, at most can be achieved 40 simultaneously The detection of index.The mode of this single sample multi objective also realizes that more pathogen screenings in face of illness mention for molecular diagnosis Having supplied may.

The full extraction molecular diagnosis micro-fluidic chip has ontology in one of the embodiments, and the ontology opens up There are storage chamber, the first capillary valves, sample cavity, Drainage pipe, the first siphon piping, the second siphon piping, the first liquid feeding chamber, second Liquid feeding chamber, the first cushion chamber, third siphon piping, collection conduits, the first controlled liquid feeding chamber, the second controlled liquid feeding chamber, the first gas Body pipeline, second gas pipeline, the first charging line, the second charging line, elution pipeline, convergence chamber, the second capillary valves, distribution Chamber, waste solution channel, collection conduit, the second cushion chamber, the 4th siphon piping, the first waste liquid chamber, sample distribution pipeline, each measurement Some or all of in chamber, each PCR amplification chamber, the second waste liquid chamber and reagent conveyance conduit；First well, the second well, First liquid filling hole, the first cushion hole, the second liquid filling hole, waste liquid venthole and venthole are each passed through the ontology and connection is external Environment.Magnetic part is equipped in sample cavity in one of the embodiments, cracking is made for cooperating external magnetic environment to realize jointly With；Alternatively, being equipped with magnetic part and abrasive article in sample cavity.Magnetic part includes magnet or magnetite, shape in one of the embodiments, Shape is unlimited；Abrasive article includes vitreum or stainless steel in one of the embodiments,；It grinds in one of the embodiments, Part is sphere, such as bead etc..Further, PCR amplification is intracavitary in one of the embodiments, presets paraffin, is used for Melt in PCR amplification and seals PCR amplification chamber.

Further, the ontology has whole closure portion in one of the embodiments, and whole closure portion is for whole Cover each chamber and its associated conduit.The whole closure portion is used for whole capped storage chamber, the in one of the embodiments, One capillary valves, sample cavity, Drainage pipe, the first siphon piping, the second siphon piping, the first liquid feeding chamber, the second liquid feeding chamber, first Cushion chamber, third siphon piping, collection conduits, the first controlled liquid feeding chamber, the second controlled liquid feeding chamber, first gas pipeline, second Gas pipeline, the first charging line, the second charging line, elution pipeline, convergence chamber, the second capillary valves, distribution chamber, sewer pipe Road, the second cushion chamber, the 4th siphon piping, the first waste liquid chamber, sample distribution pipeline, respectively measures chamber, each PCR expansion at collection conduit Increase whole presence in chamber, the second waste liquid chamber and reagent conveyance conduit.First well, second in one of the embodiments, Well, the first liquid filling hole, the first cushion hole, the second liquid filling hole, waste liquid venthole and venthole are each passed through the ontology Same side and connection external environment.

Storage chamber is Proteinase K storage chamber in one of the embodiments, for accommodating Proteinase K；First well is used It is added in storage chamber in by Proteinase K；Sample cavity is for accommodating sample to be tested, the second sample-adding in one of the embodiments Hole is for sample to be added in sample cavity；The first liquid feeding chamber is cracking neutralizer storage chamber in one of the embodiments, is used Neutralizer is cracked in accommodating, the first liquid filling hole is added in cracking neutralizer storage chamber for that will crack neutralizer；Wherein one In a embodiment, the first cushion chamber is for accommodating buffer, and the first cushion hole is for buffer to be added in the first cushion chamber； The first controlled liquid feeding chamber is as the first cleaning solution storage chamber in one of the embodiments, for accommodating the first cleaning solution, second Controlled liquid feeding chamber is as the second cleaning solution storage chamber, for accommodating the second cleaning solution；Second liquid feeding in one of the embodiments, Chamber is eluent storage chamber, and for accommodating eluent, the second liquid filling hole is for eluent to be added in the second cushion chamber；At it In middle one embodiment, convergence chamber is silica gel membrane cavity, for accommodating the pellosil of stacking；Distribute in one of the embodiments, Chamber is used to that the sample after convergence processing to be distributed in the second cushion chamber according to direction of rotation, or waste liquid is distributed to first In waste liquid chamber；The sample after convergence processing for being distributed to respectively by sample distribution pipeline in order in one of the embodiments, In a measurement chamber, extra is output in the second waste liquid chamber, and the sample in measurement chamber is under the influence of centrifugal force, defeated by reagent It send pipeline to enter in PCR amplification chamber, completes PCR amplification herein.It is understood that each pipeline by area according to demand Such as the setting such as centrifugal rotational speed.In one of the embodiments, before the use, Proteinase K, cracking neutralizer, first Cleaning solution, the second cleaning solution is preset to be encapsulated by low melt point paraffin at one end, the other end by the aluminium-foil containers of glue bond among.In advance It is equipped with Proteinase K, cracks neutralizer, the gluing end of the first cleaning solution, the aluminium-foil containers of the second cleaning solution is fixed with glue respectively again Proteinase K storage chamber, cracking neutralizer storage chamber, the first cleaning solution storage chamber, the second cleaning solution storage chamber upper area, Wherein gluing end is close to the center of circle, and paraffin sealing end is far from the center of circle.On the other hand, eluent is preset is sealed by high melting point paraffin at one end Dress, one end by the aluminium-foil containers of glue bond, the upper area of glue fixed eluant storage chamber is used at the gluing end of aluminium-foil containers again, Wherein gluing end is close to the center of circle, and paraffin sealing end is far from the center of circle.The regional temperature locating for the aluminium-foil containers rise to paraffin melting point with On, paraffin melting, the liquid being stored in aluminium-foil containers can be just released.It is blocked at first paraffin valve with high melting point paraffin First gas pipeline；Second gas pipeline is blocked with low melt point paraffin at second paraffin valve.

Cooperation below provides a full extraction molecular diagnosis micro-fluidic chip whole detection process, and details are as follows.

Protease k is added in storage chamber by the first well, and sample is added in sample cavity by the second well；

2000rpm centrifugation counterclockwise, interior temperature controlled region are warming up to 60 DEG C, are in protease k storage chamber (i.e. storage chamber), cracking Neutralizer storage chamber, the first cleaning solution storage chamber, aluminium-foil containers in the second cleaning solution storage chamber paraffin end melt, liquid examination Agent release；At the same time, the first capillary valves of protease k solution breakthrough in protease k storage chamber enter cracking chamber i.e. sample cavity In；

Centrifugal speed is down to 600rpm counterclockwise, and keeps this revolving speed 10min, while inner circle warm area maintains 60 DEG C of 10min, The cracking of magnetic agitation grinding；Micro-fluidic chip is in when full extraction molecular diagnosis micro-fluidic chip turns to the intracavitary small magnet of cracking When lower section is fixed on the top of the big magnet on instrument, due to big magnet be located closer to centrifugation the center of circle, at this time small magnet by The magnetic field force of the direction the center point arrived is greater than the centrifugal force far from the center point, and successively small magnet is moved under magnetic field force effect and split Solve the position in the intracavitary close center of circle；And when chip turns to and cracks intracavitary small magnet far from big magnet, magnetic field force is weaker very To disappearance, under the influence of centrifugal force, small magnet can come back to the bottom of cracking chamber again；So rotated in micro-fluidic chip When, magnet constantly crack it is intracavitary move up and down, cooperation cracks intracavitary preset bead, grinds to the sample for cracking intracavitary Grinding cracks solution；

Centrifugal speed is down to 200rpm counterclockwise and keeps this revolving speed 30s, cracks the mixed liquor in chamber and 11 cracking neutralize Cracking neutralizer in liquid storage chamber is distinguished under the action of capillary force, the first siphon piping and the second siphon piping inner surface thing Hydrophily processing has been first passed through, therefore with the presence of biggish capillary force, has filled up the first siphon piping and the second siphon piping；

Centrifugal speed rises to 2000rpm counterclockwise and keeps this revolving speed 1min, in the first siphon piping, the second siphon piping Liquid breaks through capillarity and enters cracking neutralization chamber, then with the help of centrifugal force, due to siphonic effect, cracks intracavitary mix The cracking neutralizer for closing liquid and cracking in neutralizer storage chamber is fully flowed into via the first siphon piping, the second siphon piping respectively In first cushion chamber；

2000rpm counterclockwise maintains 3min, and the first buffering intracavity liquid is sufficiently cracked and neutralized；

Centrifugal speed is down to 200rpm counterclockwise, rises to 2000rpm counterclockwise, the liquid in the first cushion chamber after maintaining 30s After body breaks through third siphon piping and flows through the pellosil for converging chamber, the first waste liquid chamber is entered under the action of Coriolis force, The intracorporal DNA of liquid is attracted on pellosil at this time；

2000rpm counterclockwise is maintained, outer temperature control temperature rises to 60 DEG C, No. 54 paraffin melting, paraffin valves at the first paraffin valve It opens, the first cleaning solution storage chamber is connected via first gas pipeline with the venthole of the second waste liquid chamber.First cleaning solution is able to Release, and after being cleaned to the DNA on pellosil, the first waste liquid chamber is entered under the action of Coriolis force；

2000rpm counterclockwise is maintained, outer temperature controlled region temperature rises to 80 DEG C, and the second paraffin valve is opened, and the second cleaning solution is able to Release, and after being cleaned to the DNA on pellosil, the first waste liquid chamber is entered under the action of Coriolis force；

Centrifugal speed, which is changed to, to be rotated clockwise, and speed is 500rpm clockwise；

Inner circle warm area temperature rises to 70 DEG C, the preset intracavitary aluminium-foil containers of eluent it is paraffin melting, eluent is released It puts and flows into convergence chamber, preset several layers of pellosils, due to the presence of the second capillary valves, 500rpm eluent can not be after Continuous flow down, therefore eluent is able to impregnate pellosil, 500rpm clockwise maintains 5min；

Centrifugal speed rises to 2000rpm clockwise, and the eluent after eluting silica gel film flows under the action of Coriolis force Enter in the second cushion chamber；Need to particularly point out, distribute chamber presence be in order to increase the time that Coriolis force acts on liquid, To make liquid more reliable when doing channel conversion, including switches still switch to the right to the left；

Centrifugal speed is down to 200rpm clockwise, keeps 30s, further accelerates to 1000rpm, the eluent after eluting silica gel film It breaks through the 4th siphon piping to enter sample distribution pipeline and fill up each measurement chamber one by one, extra eluent enters the second waste liquid chamber It is interior；

Centrifugal speed rises to 3000rpm clockwise, respectively measures intracavitary liquid and separately flows into each PCR amplification chamber, and dissolves It is preset at the reagent dry powder of PCR amplification chamber, in particular, PCR amplification is intracavitary to preset paraffin, paraffin is in PCR amplification Melt and seal PCR amplification chamber, to prevent the cross contamination due to caused by the escape of aerosol in amplification of 8 amplification chambers；

Centrifugal speed is down to 500rpm, and PCR temperature cycles start to expand；

It is 95 DEG C of 3min thermal startings, 95 DEG C of 15s denaturation, 60 DEG C of 40s annealing and extension that PCR, which reacts timing, does 40 temperature Circulation, starts 2 fluorescence channels in 60 DEG C of 30s and acquires fluorescence signal one by one.

In each embodiment, micro-fluidic includes but is not limited to PMMA, PDMS, PC, ABS, COC, COP etc..It is micro- The packaged type of fluidic chip includes but is not limited to hot pressing, ultrasonic bonding, laser welding, gluing etc..

A kind of full extraction molecular diagnosis micro-fluidic chip is as shown in Figure 1, it is revolved with target in one of the embodiments, Turn center and is located at except the full top for extracting molecular diagnosis micro-fluidic chip, the full extraction molecular diagnosis micro-fluidic chip packet It is controlled to include storage chamber 102, sample cavity 105, the first liquid feeding chamber 111, the first cushion chamber 113, the first controlled liquid feeding chamber 116, second Liquid feeding chamber 117, convergence chamber 124, the second liquid feeding chamber 110, distribution chamber 126, the first waste liquid chamber 132, cushion chamber 129, second are useless Sap cavity 139, at least one measurement chamber 134 and an at least PCR amplification chamber 135；Also referring to Fig. 3, Fig. 4 and Fig. 5, press and the mesh Mark rotation center distance from small to large sequence arrangement are as follows: storage chamber 102, sample cavity 105, the first cushion chamber 113, first by Control liquid feeding chamber 116, the second controlled liquid feeding chamber 117, convergence chamber 124, distribution chamber 126, the first waste liquid chamber 132, measurement chamber 134, PCR amplification chamber 135；First liquid feeding chamber 111 be greater than at a distance from target rotation center storage chamber 102 and target rotation center away from From and less than the first cushion chamber 113 at a distance from target rotation center；Second liquid feeding chamber 110 is big at a distance from target rotation center In the first cushion chamber 113 at a distance from target rotation center and be less than convergence chamber 124 at a distance from target rotation center；Cushion chamber 129 are more than or equal to the first waste liquid chamber 132 at a distance from target rotation center at a distance from target rotation center and are less than measurement chamber 134 at a distance from target rotation center；Second waste liquid chamber 139 is greater than cushion chamber 129 and target at a distance from target rotation center The distance of rotation center；Storage chamber 102 is equipped with the first well 101 at close-target rotation center, and storage chamber 102 passes through first Capillary valves 103 are connected to sample cavity 105；Sample cavity 105 is equipped with the second well 104, sample cavity 105 at close-target rotation center The first cushion chamber 113 is connected to by the first siphon piping 107；First liquid feeding chamber 111 is equipped with first at close-target rotation center Liquid filling hole 109, the first liquid feeding chamber 111 are connected to the first cushion chamber 113 by the second siphon piping 108；First cushion chamber 113 is close The first cushion hole 112 is equipped at target rotation center, the first cushion chamber 113 passes through the connection convergence chamber of third siphon piping 114 124；Second liquid feeding chamber 110 is equipped with the second liquid filling hole 122 at close-target rotation center, and the second liquid feeding chamber 110 passes through elution pipe The connection of road 123 convergence chamber 124；First controlled liquid feeding chamber 116 is connected at close-target rotation center by first gas pipeline 118 The first paraffin valve 137 is equipped in second waste liquid chamber 139 and first gas pipeline 118, the first controlled liquid feeding chamber 116 adds by first The connection of liquid pipe road 120 convergence chamber 124；Second controlled liquid feeding chamber 117 passes through second gas pipeline 119 at close-target rotation center It is connected in the second waste liquid chamber 139 and second gas pipeline 119 and is equipped with the second paraffin valve 136, the second controlled liquid feeding chamber 117 passes through the Two charging lines 121 connection convergence chamber 124；It converges chamber 124 and chamber 126 is distributed by the connection of the second capillary valves 125, it is described to mention entirely Molecular diagnosis micro-fluidic chip is taken to be equipped with filter structure in convergence chamber 124；Wherein, the first cushion chamber 113 passes through third siphon Pipeline 114 be connected to collection conduits 115, the second liquid feeding chamber 110 by elution pipeline 123 be connected to collection conduits 115, first it is controlled plus Sap cavity 116 is connected to collection conduits 115 by the first charging line 120, and the second controlled liquid feeding chamber 117 passes through the second charging line 121 connection collection conduits 115；The connection convergence chamber 124 of collection conduits 115.Distribute chamber 126 far from one at target rotation center End is connected to the first waste liquid chamber 132 by waste solution channel 127, and the other end is connected to the second cushion chamber 129 by collection conduit 128, and Waste solution channel 127 and collection conduit 128 are located at the two sides of distribution chamber 126；First waste liquid chamber 132 is in close-target rotation Waste liquid venthole 131 is equipped at the heart；Second cushion chamber 129 is connected to sample by the 4th siphon piping 130 and distributes pipeline 133；Sample 133 sequential communication of this distribution pipeline respectively measures chamber 134, and the end of sample distribution pipeline 133 passes through the connection of Drainage pipe 106 the Two waste liquid chambers 139；Measurement chamber 134 is identical as the quantity of PCR amplification chamber 135 and is correspondingly arranged, each connection of measurement chamber 134 one PCR amplification chamber 135；Also, the full extraction molecular diagnosis micro-fluidic chip further includes reagent conveyance conduit 140, measures chamber 134 Identical as the quantity of reagent conveyance conduit 140 and be correspondingly arranged, each measurement chamber 134 passes through corresponding reagent conveyance conduit The corresponding PCR amplification chamber 135 of 140 connections.Second waste liquid chamber 139 is equipped with venthole 138 at close-target rotation center.It can be with Understand, in embodiment illustrated in fig. 1, each chamber be it is closed, be similar to the structure of lid with one to close these chambers Room and its pipeline only reserve a some holes, as shown in Fig. 2, the first well 101, the second well 104, the first liquid filling hole 109, First cushion hole 112, the second liquid filling hole 122, waste liquid venthole 131 and venthole 138 are each passed through the same side of the ontology Face and connection external environment.

As shown in Figure 1, the ontology has partial sector structure, it is corresponding that target rotation center is located at the partial sector structure The center point；Alternatively, target rotation center is located between the corresponding center of circle of partial sector structure and the top of the ontology.

In order to enhance siphonage, each siphon piping, including the first siphon piping, second in one of the embodiments, Siphon piping, third siphon piping and the 4th siphon piping, inner surface have hydrophilic layer or pass through hydrophilic treated.Such as Fig. 1 institute Show, the 4th siphon piping 130 has liquid input section 231, ascent stage 232, top changeover portion 233 and descending branch 234, liquid input section 231 For inputting liquid, ascent stage 232, top changeover portion 233 and descending branch 234 cooperatively form similar П font or a ∩ font Structure, to form a siphon piping and play the role of siphon valve；Such syphon structure design, in centrifugal force very little (low-speed centrifugal) or do not have centrifugal force (stopping operating) when, the chamber indoor liquid by close-target rotation center is by capillary force Traction does not cross the siphon piping with siphonage at close-target rotation center, until hydraulically full in siphon piping；With After increase centrifugal speed, under the influence of centrifugal force, siphon flowing occurs inside siphon piping, and liquid all flows into separate below In the chamber of target rotation center.

In the application one embodiment, a kind of microfluidic system comprising extract molecule described in any embodiment entirely and examine Disconnected micro-fluidic chip.Above-mentioned microfluidic system exists the nucleic acid of the equivalent distribution of sample cracking, nucleic acid purification, reagent, multi-chamber The all processes such as the lower amplification of specific primer constraint are all integrated in a centrifugal microfluidic control chip, and thus, it is possible to realize that fluorescence is believed Number acquisition and analysis etc. subsequent analysis work, realize the diagnostic mode of single sample multi objective, also be molecular diagnosis realize face Possibility is provided to more pathogen screenings of illness；Also, entire reaction process is in closed micro-fluidic chip, is reduced A possibility that burden and pollution of operator, but also entire molecular diagnostic procedure is no longer dependent on molecular diagnostic laboratories, Also it is no longer dependent on professional operator, realizes the demand quickly detected whenever and wherever possible, is medical inspection and control and prevention of disease Bring huge help.

The concrete application for providing microfluidic system or its full extraction molecular diagnosis micro-fluidic chip is continued with, mammitis is One of most common disease of milk cow, and a kind of maximum disease is endangered to milk cow production.Mammitis refer to cow mammary gland by A kind of inflammatory pathological change occurred to stimulations such as physics, chemistry and microorganisms, its main feature is that physicochemical property has occurred in milk Variation, quantity of leucocyte increases and the pathological change of breast tissue.Mammitis is divided into clinic mastitis and recessive breast It is two kinds scorching.Chinese milk cow clinic mastitis disease incidence is 33.41% (9.7%~55.6%), recessive mastitis average positive Rate is 73.91%, and newborn room illness rate is 44.74%.Mastitis for milk cows not only causes milk production of cow decline, milk-quality to change, Even can the area Zhi Ru suppurate, gangrene, lose the ability of lactation, cause huge economic losses.Simultaneously because clinically using a large amount of Antibiotics, there are the risks of medicament residue, affect the quality of cow's milk.Therefore mastitis for milk cows prevention and treatment increasingly by To the attention of herding worker.Garget detects micro-fluidic consumptive material and cooperates full-automatic foranalysis of nucleic acids system, can be at about 70 minutes It is interior to single milk sample provide up to 16 indexs testing result (due on micro-fluidic chip there are 8 PCR amplification chambers, Therefore 16 indexs only need two fluorescence channels).

The process flow of the reagent of garget detection is described as follows:

20 μ L Proteinase Ks (20mg/ml) are added in storage chamber；

200 μ L milk samples are added in sample cavity；

200 μ L Lysis Buffer are added in sample cavity as lysate；

200 μ L dehydrated alcohols are added in the first liquid feeding chamber as neutralizer；

500 μ L Wash Buffer are added in the first controlled liquid feeding chamber as the first cleaning solution；

500 μ L Wash Buffer are added in the second controlled liquid feeding chamber as the second cleaning solution；

100 μ L Elution Buffer are added in the second liquid feeding chamber as eluent；

Each group PCR amplification material is added separately in each PCR amplification chamber；

Each Step Time is determined by traditional cow room inflammation detection mode, using related embodiment above and carries out centrifugation control It is controlled with temperature, using simple, conveniently, realizes the demand quickly detected whenever and wherever possible, entire reaction process is in close In the micro-fluidic chip closed, a possibility that reducing the burden and pollution of operator.

It should be noted that the other embodiments of the application further include, the mutually group of the technical characteristic in the various embodiments described above Close be formed by, the full extraction molecular diagnosis micro-fluidic chip that can implement and microfluidic system.

Each technical characteristic of above embodiments can be combined arbitrarily, for simplicity of description, not to above-described embodiment In each technical characteristic it is all possible combination be all described, as long as however, the combination of these technical characteristics be not present lance Shield all should be considered as described in this specification.

Above embodiments only express the several embodiments of the application, and the description thereof is more specific and detailed, but can not Therefore it is interpreted as the limitation to claim.It should be pointed out that for those of ordinary skill in the art, In Under the premise of not departing from the application design, various modifications and improvements can be made, these belong to the protection scope of the application. Therefore, the scope of patent protection of the application should be determined by the appended claims.

Claims

1. a kind of complete extract molecular diagnosis micro-fluidic chip, with target rotation center, which is characterized in that including storage chamber, Sample cavity, the first liquid feeding chamber, the first cushion chamber, at least one controlled liquid feeding chamber, the second liquid feeding chamber, distribution chamber, the first waste liquid chamber, Cushion chamber, the second waste liquid chamber, at least one measurement chamber and an at least PCR amplification chamber；

By at a distance from the target rotation center from small to large sequence arrangement are as follows: storage chamber, sample cavity, the first cushion chamber, Controlled liquid feeding chamber, distribution chamber, the first waste liquid chamber, measurement chamber, PCR amplification chamber；

It is at a distance from target rotation center and slow less than first that first liquid feeding chamber is greater than storage chamber at a distance from target rotation center Chamber is rushed at a distance from target rotation center；

Second liquid feeding chamber is greater than the first cushion chamber at a distance from target rotation center and is less than at a distance from target rotation center and divided Chamber is sent out at a distance from target rotation center；

Cushion chamber, which is more than or equal to the first waste liquid chamber at a distance from target rotation center and is less than at a distance from target rotation center, to be surveyed Chamber is measured at a distance from target rotation center；

Storage chamber is equipped with the first well at close-target rotation center, and storage chamber is connected to sample cavity by the first capillary valves；

Sample cavity is equipped with the second well at close-target rotation center, and sample cavity passes through the first buffering of the first siphon piping connection Chamber；

First liquid feeding chamber is equipped with the first liquid filling hole at close-target rotation center, and the first liquid feeding chamber is connected to by the second siphon piping First cushion chamber；

First cushion chamber is equipped with the first cushion hole at close-target rotation center, and the first cushion chamber is connected to by third siphon piping Distribute chamber；

Second liquid feeding chamber is equipped with the second liquid filling hole at close-target rotation center, and the second liquid feeding chamber passes through elution pipeline connection distribution Chamber；

Controlled liquid feeding chamber is connected in the second waste liquid chamber and gas pipeline by gas pipeline at close-target rotation center and is equipped with phase Become valve, controlled liquid feeding chamber is connected to distribution chamber by charging line；

The full extraction molecular diagnosis micro-fluidic chip is in distribution chamber or distributes the inlet of chamber or distributes entering for chamber Filter structure is equipped with before mouthful；

Distribute chamber and the first waste liquid chamber is connected to by waste solution channel far from one end at target rotation center, the other end passes through collection Pipeline is connected to the second cushion chamber, and waste solution channel and collection conduit are located at the two sides of distribution chamber；

Sample distribution pipeline sequential communication respectively measures chamber and end is connected to the second waste liquid chamber；Measure the quantity of chamber and PCR amplification chamber Identical and be correspondingly arranged, each measurement chamber is connected to a PCR amplification chamber；

2. extracting molecular diagnosis micro-fluidic chip entirely according to claim 1, which is characterized in that the full extraction molecular diagnosis Micro-fluidic chip includes two controlled liquid feeding chambers, the respectively first controlled liquid feeding chamber and the second controlled liquid feeding chamber；

By at a distance from the target rotation center from small to large sequence arrangement are as follows: storage chamber, sample cavity, the first cushion chamber, First controlled liquid feeding chamber, the second controlled liquid feeding chamber, distribution chamber, the first waste liquid chamber, measurement chamber, PCR amplification chamber；

First controlled liquid feeding chamber is connected to the second waste liquid chamber and first gas by first gas pipeline at close-target rotation center The first paraffin valve is equipped in pipeline, the first controlled liquid feeding chamber passes through the first charging line connection distribution chamber；

Second controlled liquid feeding chamber is connected to the second waste liquid chamber and second gas by second gas pipeline at close-target rotation center The second paraffin valve is equipped in pipeline, the second controlled liquid feeding chamber passes through the second charging line connection distribution chamber.

3. according to claim 1 entirely extract molecular diagnosis micro-fluidic chip, which is characterized in that further include convergence chamber, by with The sequence arrangement of the distance of the target rotation center from small to large are as follows: controlled liquid feeding chamber, convergence chamber, distribution chamber；

Second liquid feeding chamber, which is greater than the first cushion chamber at a distance from target rotation center and is less than at a distance from target rotation center, to converge Poly- chamber is at a distance from target rotation center；

4. extracting molecular diagnosis micro-fluidic chip entirely according to claim 3, which is characterized in that the first cushion chamber passes through third Siphon piping is connected to collection conduits, and the second liquid feeding chamber is connected to collection conduits by elution pipeline, and controlled liquid feeding chamber passes through liquid-feeding tube Road is connected to collection conduits, collection conduits connection convergence chamber.

5. according to the full extraction molecular diagnosis micro-fluidic chip of claim 3 or 4, which is characterized in that be equipped in convergence chamber Filter membrane is as the filter structure；Further, the filter membrane is the multilayer pellosil being stacked.

6. extracting molecular diagnosis micro-fluidic chip entirely according to claim 3, which is characterized in that the full extraction molecular diagnosis Micro-fluidic chip includes two controlled liquid feeding chambers, the respectively first controlled liquid feeding chamber and the second controlled liquid feeding chamber；

By at a distance from the target rotation center from small to large sequence arrangement are as follows: storage chamber, sample cavity, the first cushion chamber, First controlled liquid feeding chamber, the second controlled liquid feeding chamber, convergence chamber, distribution chamber, the first waste liquid chamber, measurement chamber, PCR amplification chamber；

First controlled liquid feeding chamber is connected to the second waste liquid chamber and first gas by first gas pipeline at close-target rotation center The first paraffin valve is equipped in pipeline, the first controlled liquid feeding chamber passes through the first charging line connection convergence chamber；

Second controlled liquid feeding chamber is connected to the second waste liquid chamber and second gas by second gas pipeline at close-target rotation center The second paraffin valve is equipped in pipeline, the second controlled liquid feeding chamber passes through the second charging line connection convergence chamber；

Further, the first cushion chamber is connected to collection conduits by third siphon piping, and the second liquid feeding chamber is connected by elution pipeline Logical collection conduits, the first controlled liquid feeding chamber are connected to collection conduits by the first charging line, and the second controlled liquid feeding chamber passes through second Charging line is connected to collection conduits；Collection conduits connection convergence chamber.

7. extracting molecular diagnosis micro-fluidic chip entirely according to claim 1, which is characterized in that further include reagent delivery pipe Road, measurement chamber is identical as the quantity of reagent conveyance conduit and is correspondingly arranged, and each measurement chamber passes through corresponding reagent delivery pipe Road is connected to corresponding PCR amplification chamber；Alternatively, the end of sample distribution pipeline is connected to the second waste liquid chamber by Drainage pipe.

8. according to claim 1 to molecular diagnosis micro-fluidic chip is extracted described in any one of 7 entirely, which is characterized in that described complete Extracting molecular diagnosis micro-fluidic chip has ontology, and the ontology offers storage chamber, the first capillary valves, sample cavity, drain pipe Road, the second siphon piping, the first liquid feeding chamber, the second liquid feeding chamber, the first cushion chamber, third siphon piping, converges at the first siphon piping Collect pipeline, the first controlled liquid feeding chamber, the second controlled liquid feeding chamber, first gas pipeline, second gas pipeline, the first charging line, Second charging line, elution pipeline, convergence chamber, the second capillary valves, distribution chamber, waste solution channel, collection conduit, the second buffering Chamber, the first waste liquid chamber, sample distribution pipeline, respectively measures chamber, each PCR amplification chamber, the second waste liquid chamber and reagent at the 4th siphon piping Some or all of in conveyance conduit；First well, the second well, the first liquid filling hole, the first cushion hole, the second liquid feeding Hole, waste liquid venthole and venthole are each passed through the ontology and connection external environment；

And/or magnetic part and/or abrasive article are equipped in sample cavity；

And/or PCR amplification is intracavitary presets paraffin.

9. extracting molecular diagnosis micro-fluidic chip entirely according to claim 8, which is characterized in that the first well, second add Sample hole, the first liquid filling hole, the first cushion hole, the second liquid filling hole, waste liquid venthole and venthole are each passed through the same of the ontology One side and connection external environment.

10. a kind of microfluidic system, which is characterized in that including extracting molecular diagnosis entirely as described in any one of claims 1 to 9 Micro-fluidic chip.