CN207793229U - Molecule diagnoses micro-fluidic chip and molecule diagnoses micro-fluidic chip system - Google Patents

Abstract

The utility model is related to micro-fluidic chip molecular diagnosis field, more particularly to molecule diagnosis micro-fluidic chip and molecule diagnoses micro-fluidic chip system.Chip includes：The sample injection unit (1) for receiving sample to be checked, the liquid storage unit (2) for storing reaction reagent, the cell cracking unit (3) that cell is cracked, the nucleic acid amplification unit (4) that nucleic acid is expanded；Cell cracking unit includes that slit (31), the first chamber (32) separated by slit and second chamber (33), at least one chamber are configured with grinding microballon；First chamber is communicated with sample injection unit, and second chamber is communicated at least one liquid storage unit, receives cell pyrolysis liquid.Chip system is by upper, middle and lower up of three layers；Middle level is the chip；Levels are for covering closing middle level；There are the sample holes being connect with sample injection unit, the relief hole of corresponding liquid storage unit in upper layer.The chip detection process is simple, can be quantified to reaction sample, high sensitivity, repeatability are strong.

Description

Molecule diagnoses micro-fluidic chip and molecule diagnoses micro-fluidic chip system

Technical field

It is the utility model is related to micro-fluidic chip molecular diagnosis field, more particularly to a kind of for the micro-fluidic of molecule diagnosis Chip and a kind of fluidic chip system for molecule diagnosis.

Background technology

Microfluidic chip technology (microfluidics) is otherwise known as Microfluid based Lab on a chip or chip lab (lab-on-a-chip), the chemistry or biology laboratory that are built on one piece several square centimeters of chip are referred to, it is chemistry It with sample preparation involved in the fields such as biology, reacts, detaches, detection, cell culture, sorting, the basic operations such as cracking Unit is integrated on the chip of one piece of very little, and network is formed by microchannel, runs through whole system with controlled fluid, to realize life Object, chemical, the various functions in the fields such as medical diagnosis.

The essential characteristic and sharpest edges of microfluidic chip technology be：A variety of cellular constructions can on small chip platform With flexible combination so that chip design is upper flexible and changeable and multiple functional；The small required detection of chip internal structure unit Sample size is considerably less, and the bigger serface of microstructure unit allows interior reagent quickly to spread to realize fast reaction and inspection It surveys；Micro-fluidic chip is usually automatically performed internal-response by necessary instrument, so during actual test, micro-fluidic chip Technology can reduce the technology requirement to medicine testing staff, reduce the human error of detection, and then reduce the medicine inspection of patient Survey cost；Microfluidic chip technology using automation equipment due to being completed, so can be with so internal-response process is fully controllable It is more accurate to obtain, more sensitive detection data.

In medical science, in-vitro diagnosis (In-Vitro-Diagnostics, IVD) is one very important point Branch, refers to being detected to obtain to the sample (such as blood, urine, the body fluid such as saliva) from human body in human vitronectin Clinical diagnosis information, according to the information come judge subject whether with certain disease or be subjected to certain germ infection diagnosis side Method.Currently, with biotechnology, especially biochemistry, immunology, the fast development of molecular biology, in-vitro diagnosis according to Testing principle and detection method mainly have biochemical diagnosis, immunodiagnosis, molecule diagnosis, microbe diagnosis, the diagnosis of blood coagulation class, streaming Cyto-diagnosis etc., mesophytization are immunized, and molecule diagnosis is the major component field of current China's in-vitro diagnosis.

Molecule diagnosis be using molecular biology technology and methods research human endogenous property or exogenous biomolecule with The presence of molecular system, structure or expression regulation variation, for the prevention of disease, prediction, diagnosis, treat and lapse to provide information and Decision-making foundation.High sensitivity that molecular diagnostic techniques have with it, high specificity, it is easy quickly, qualitative and quantitative analysis can be carried out Etc. advantages, be widely used in disease forecasting, prevent, prognosis, Individual drug treatment, Prenatal Screening, tumour, heredity Disease, blood disease, infectious diseases (including bacterial infection, viral infection, parasite, fungi, zoonosis etc.), god Through multiple fields such as psychotic disorder, organ transplant, inborn defects, inspection result to the assessment of body neurological susceptibility, medical diagnosis on disease, Prognosis, Treatment monitoring, Individual drug treatment, genetic counselling, health control and household fertility plan etc. all have weight The reference value wanted even decisive value.

The molecular diagnostic techniques platform that China routinely carries out at present mainly has flow cytometry, molecular hybridization, DNA/ RNA sequencing technologies and nucleic acid amplification technologies.But the application field of flow cytometry is narrow, is only applied at present to circulating tumor The liquid biopsy field that cell is sorted, it is then helpless for the context of detection of infectious disease.Molecular hybridization can only be It is carried out in special detection section office, it is complicated for operation, it is easy, by environmental pollution, to need efficient staff to operate.DNA/RNA is surveyed Sequence technology needs expensive mating sequencing instrument, and sequencing is of high cost, and time-consuming.Nucleic acid amplification technologies are in molecular diagnosis field Application range is most wide, and easy to operate, it is only necessary to which detection can be completed in PCR amplification instrument, and the manual operation needed is less.

Nevertheless, the patent for carrying out nucleic acid amplification molecule diagnosis using microfluidic chip technology is especially few, both at home and abroad will The company that both technologies combined and then developed the chip product of maturation is less.It investigates and finds by patent, both at home and abroad in recent years To have one is disclosed using the related patents of microflow control technique progress molecule diagnosis, such as Chinese patent 201510265559 on a small quantity The kind equidistant decile nucleic acid amplification micro-fluidic chip of high integration, but the chip only has the function of nucleic acid amplification, without reagent Storage, reagent mixing, the functions such as sample amounts；A kind of centrifugal disk is disclosed in Chinese patent 201610242532 for another example Array detection of nucleic acids micro-fluidic chip, but the chip is detected just for nucleic acid extraction liquid, needs user before use Sample to be checked is first subjected to the nucleic acid in processing extraction sample by hand, then nucleic acid is injected in the chip again and is detected, it should Chip improves user's requirement in use, needs in advance to come out nucleic acid extraction.

Utility model content

The purpose of this utility model is to overcome problem as above of the existing technology, provide a kind of and nucleic acid extraction and The micro-fluidic chip for the diagnosis of nucleic acid amplification molecule of sample amounts, the micro-fluidic chip have detection process simple, can Reaction sample is quantified, and nucleic acid extraction is carried out to reaction sample, and micro-fluidic chip provided by the utility model is each Structural unit is interconnected, and detection sensitivity is high, repeatability is strong.

To achieve the goals above, on the one hand the utility model provides a kind of molecule diagnosis micro-fluidic chip, this is micro-fluidic Chip includes：For receiving the sample injection unit of sample to be checked, the liquid storage unit for storing reaction reagent, for treating sample sheet In cell the cell cracking unit, the nucleic acid amplification unit for being expanded to nucleic acid that are cracked；

Wherein, the cell cracking unit includes slit and the first chamber and second chamber that are separated by slit, described At least one of first chamber and the second chamber chamber are configured with grinding microballon；The first chamber and the sample introduction list Member communicates, and the second chamber is communicated at least one liquid storage unit, for receiving cell pyrolysis liquid.

Preferably, the cell cracking unit further includes baffle, and the baffle is opposite with the slit but does not connect, described First chamber and second chamber are communicated by the baffle with the space that the slit defines.

Preferably, the micro-fluidic chip further includes being arranged between the cell cracking unit and nucleic acid amplification unit Nucleic acid purification unit, to be carried out to the pyrolysis product of the cell cracking unit before nucleic acid enters the nucleic acid amplification unit It purifies, configured with the magnetic bead for capableing of specific adsorption nucleic acid in the nucleic acid purification unit.

Preferably, the nucleic acid amplification unit includes being fixed at least in micro- well by the array of multiple micro- well constructions A kind of primer pair.

Preferably, the micro-fluidic chip further includes the waste unit for collecting waste liquid caused by each step reaction.

Preferably, which further includes the ventilation unit being connect with the waste unit, for being micro-fluidic core Piece system provides required external pressure.

Preferably, which further includes the tool interface system being connect with the ventilation unit, and the tool interface system is used Necessary instrument outside connection micro-fluidic chip system, the air pressure that the necessary instrument provides are provided to by the ventilation unit The micro-fluidic chip system.

Preferably, the necessary instrument include for provide the promotion pulsometer of air pressure, gas-guide tube and with the ventilation The air-tightness interface of unit connection；The air-tightness interface is tubaeform.

Preferably, the reaction reagent is loaded by reagent pouch in the liquid storage unit；The reagent pouch includes Location hole for being fixed in the liquid storage unit, seal and described for opening and discharging by the seal The crush-zone or needling structure of reaction reagent.

The second aspect of the utility model also provides a kind of molecule diagnosis micro-fluidic chip system, the micro-fluidic chip system It is made of the upper, middle and lower；

Wherein, media layer damage is that molecule as described above diagnoses micro-fluidic chip；

Wherein, superstructure and understructure close the middle level for covering；It is provided in superstructure and sample introduction list Member connection sample holes, and with the relief hole corresponding to liquid storage unit, for providing external impetus to discharge in liquid storage unit Reaction reagent.

The utility model can obtain following advantageous effect：

1, the utility model detects sample to be checked using microfluidic chip technology combination nucleic acid amplification, has detection sensitive The advantages that degree is high, and detection limit is low, and detection is repeatable, while coordinating particular detection instrument, full-automatic chip detection, nothing may be implemented Need human interference that can quickly obtain accurate testing result.

2, the utility model uses microfluidic chip technology, treats sample and originally carries out volume quantitative first so that only spy The sample for determining volume reacts, and ensures the accuracy of testing result, simultaneously because chip structure is fixed, so for same This its detection repeatability is strong, ensures the stability and reliability of testing result.

3, the utility model uses microfluidic chip technology, it would be desirable to participate in all liq reagent of molecule diagnostic reaction all It is pre-stored in liquid storage unit, it is entirely avoided the pollution problem in conventional open molecule diagnostic platform, while totally enclosed type liquid Body reagent, which preserves form, can extend the shelf-life of chip, it is ensured that chip remains able to be stablized for a comparatively long period of time And accurate testing result.

4, the utility model uses microfluidic chip technology, only carries out one-time detection for each detection sample, detects Chip is discarded immediately after completion, it is entirely avoided the detection interference in hospital between different samples, while patient is avoided completely Between cross-infection.

5, the utility model uses microfluidic chip technology, and the waste liquid and extra sample after reaction is completed are all completely close It is enclosed in chip interior, the leakage of waste liquid or sample will not be caused, it is nuisanceless to detection environment, to hospital's testing staff's safety Height will not lead to the generation of nosocomial infection.

6, micro-fluidic chip provided by the utility model can note user by the mutual cooperation of above-mentioned each structural unit The sample to be checked entered carries out accurate quantification, is sufficiently mixed so that the bacterium contained in sample, virus or other pathogen cells It carries out rapid cleavage and releases internal nucleic acid substances, high quality is further preferably obtained by isolating and purifying for silica gel magnetic bead DNA/RNA molecules can be detected the DNA/RNA molecules of specific pathogen bacterium using specific nucleic acid amplification method such as PCR etc., To detect in sample to be checked whether to include certain pathogen, accurate " positive " or " feminine gender " result is obtained.

Description of the drawings

Fig. 1 is a kind of specific molecule diagnosis micro-fluidic chip schematic diagram provided by the utility model.

Fig. 2 shows a kind of specific necessary instrument and its connection type with molecule diagnosis micro-fluidic chip.

Fig. 3 is the structure and its Operational Mechanisms figure of cell cracking unit provided by the utility model；Wherein, the utility model Shown in structure such as Fig. 3 (A) of the cell cracking unit of offer；Fig. 3 (B) shows that when being detected micro-valve door is opened, to be checked Sample is entered by sample inlet in first chamber 32, and extra sample can enter second chamber 33 by slit 31, and It is flowed out by micro-valve door；Fig. 3 (C) is shown before cell cracking operation, is first closed micro-valve door, is then passed through lysate Lysate is discharged into second chamber 33 by entrance；If Fig. 3 (D) shows that two milling chambers are squeezed in succession, to promote So that lysate and sample to be checked is mixed, is operated by the alternating of two pressure rams, mixed liquor can be squeezed from a chamber Enter another chamber quickly through slit 31, while under the action of grinding microballon, cell membrane or cell in sample to be checked Rapid cleavage occurs for wall, and the function of cell cracking is realized with this.

Fig. 4 (A) is a kind of schematic diagram of reagent pouch provided by the utility model；Fig. 4 (B), which is shown, passes through external pressure The mode of disrupting agent pouch；Fig. 4 (C) is shown in such a way that needle pierces disrupting agent pouch.

Fig. 5 is that the molecule of the utility model diagnoses the schematic diagram of a layer structure of micro-fluidic chip system.

Fig. 6 is a kind of schematic diagram of superstructure provided by the utility model.

Reference sign

1 sample injection unit, 2 liquid storage unit, 3 cell cracking unit

4 nucleic acid amplification unit, 5 nucleic acid purification unit, 6 waste unit

7 ventilation unit, 8 tool interface system, 9 necessary instrument

21 reagent pouch, 31 slit, 32 first chamber

33 second chamber, 34 baffle, 41 micro- well

91 pulsometer, 92 gas-guide tube, 93 air-tightness interface

211 location hole, 212 seal, 213 crush-zone

214 needling structures

Specific implementation mode

The endpoint of disclosed range and any value are not limited to the accurate range or value herein, these ranges or Value should be understood as comprising the value close to these ranges or value.For numberical range, between the endpoint value of each range, respectively It can be combined with each other between the endpoint value of a range and individual point value, and individually between point value and obtain one or more New numberical range, these numberical ranges should be considered as specific open herein.

As shown in figures 1 and 3, the utility model first aspect provides a kind of molecule diagnosis micro-fluidic chip, this is micro-fluidic Chip includes：For receiving the sample injection unit 1 of sample to be checked, the liquid storage unit 2 for storing reaction reagent, for treating sample Cell cracking unit 3, the nucleic acid amplification unit 4 for being expanded to nucleic acid that cell in this is cracked；

Wherein, the cell cracking unit 3 includes slit 31 and the first chamber 32 separated by slit 31 and the second chamber Room 33, at least one of the first chamber 32 and the second chamber 33 chamber are configured with grinding microballon；First chamber Room 32 is communicated with the sample injection unit 1, and the second chamber 33 is communicated at least one liquid storage unit 2, is split for receiving cell Solve liquid.

According to the utility model, the grinding microballon is preferably positioned over cell cracking during micro-fluidic chip encapsulates In the first chamber 32 and/or second chamber 33 of unit 3, effect is mainly ground the cell of nucleic acid to be extracted, and And the cell membrane for the cell (for example, virus or other pathogens) that shearing force is destroyed in sample to be checked is provided in process of lapping Or cell wall, so that it is released internal nucleic acid substances.Its material can be glass, garnet, silicon carbide, steel ball, ceramics etc., Diameter is 0.05mm-10mm.

According to the utility model, the effect of the slit 31 is to provide a certain amount of spatial volume, and sample is waited for excessive This is quantified, and liquid is promoted sick in sample to be checked by speed when only retaining the sample of designated volume, and can speed up grinding The rapid cleavage of opportunistic pathogen.Its slit width can be 0.001mm-10mm.It can thus be seen that cell provided by the utility model Cracking unit 3 can not only treat sample this cell and be cracked, and also have the function of quantitative.

According to the utility model, the cell cracking unit 3 is further preferably connected with necessary instrument 9, and the necessary instrument 9 wraps Push-pull rod is included, therefore, when the push-pull rod of necessary instrument 9 pushes first chamber 32, liquid can be reached quickly through slit 31 Second chamber 33, when the push-pull rod in necessary instrument pushes second chamber 33, liquid can reach first quickly through slit 31 Chamber 32, such sample to be checked can concussion moves back and forth in two milling chambers, while coordinating the grinding repeatedly of grinding bead, can To accelerate the cracking of pathogen.

According to a kind of preferred embodiment of the utility model, the cell cracking unit 3 further includes baffle 34, described Baffle 34 makes the first chamber 32 and second chamber 33 communicate with the access that the slit 31 defines.In the preferred situation Under, during cell cracking, flow velocity of the sample to be checked between first chamber 32 and second chamber 33 can be further exacerbated by And oscillation degree, to further speed up the cracking of sample cell to be checked.

The structure and operation principle of cell cracking unit 3 provided by the utility model are described in detail in conjunction with Fig. 3, As shown in Fig. 3 (A), cell cracking unit 3 includes multiple grinding microballons, the slit 31, (first chamber 32 of first chamber 32 It is connected to sample injection unit 1 by sample inlet), (second chamber 33 is equipped with lysate to second chamber 33 by the connection of lysate entrance Liquid storage unit 2), micro-valve door.Wherein grinding microballon is added in the process of chip packaging in first chamber 32, and is stored in this In chamber structure.When being detected, as shown in Fig. 3 (B), micro-valve door is opened, and sample to be checked is entered by sample inlet Into first chamber 32, extra sample can enter second chamber 33 by slit 31, and be flowed out by micro-valve door, preferably It flows into waste unit 6 mentioned below, so structure herein can also play the work for treating sample this progress volume quantitative With.As shown in Fig. 3 (C), before cell cracking operation, first micro-valve door is closed, then will be split by lysate entrance Solution liquid is discharged into second chamber 33.As shown in Fig. 3 (D), the work of the push-pull rod promotion of corresponding position in necessary instrument 9 Under, two milling chambers can be squeezed in succession, to promote lysate and sample to be checked to be mixed, be squeezed by two The alternating of bar operates, and can squeeze mixed liquor and enter another chamber quickly through slit 31 from a chamber, while grind Under the action of microballon, rapid cleavage occurs for cell membrane or cell wall in sample to be checked, and the function of cell cracking is realized with this.

Wherein, the liquid for being cracked to cell that the lysate can be known in the art, art technology Personnel can select lysate according to actual conditions.One example of the lysate is：The dodecane of 55mmol/L Basic ring acid sodium (SDS), the ethylenediamine tetra-acetic acid (EDTA) of 25mmol/L, the trishydroxymethylaminomethane of 100mmol/L (Tris), the sodium chloride (NaCl) of 500mmol/L finally adjusts the pH value of above-mentioned solution to 8.0 with NaOH solution.

According to the utility model, in order to further increase the detection result of the micro-fluidic chip, the micro-fluidic chip Further include the nucleic acid purification unit 5 being arranged between the cell cracking unit 3 and nucleic acid amplification unit 4, to enter in nucleic acid The pyrolysis product of the cell cracking unit 3 is purified before the nucleic acid amplification unit 4.Wherein, pure in the nucleic acid Change in unit 5 configured with the magnetic bead for capableing of specific adsorption nucleic acid (RNA or DNA).

According to a kind of preferred embodiment of the utility model, the magnetic bead is silica gel magnetic bead, and the silica gel magnetic bead can be with For the silica gel magnetic bead of any commercially available (for example, Dynabeads magnetic beads purchased from ThermoFisher companies).Wherein, silica gel Magnetic bead can be previously positioned in nucleic acid purification unit 5 in chip package, and the layer of silica gel on surface being capable of specific adsorption core Then acid molecule is fixed silica gel magnetic bead by the sucking action of magnet, then can will be unadsorbed by the flushing of cleaning solution The impurity such as albumen, cell membrane wash, to play the purpose of purification of nucleic acid.Nucleic acid purification unit 5 is silica gel magnetic bead and thin The place and cleaning solution that cellular lysate product is mixed rinse unadsorbed impurity place and eluent by nucleic acid molecules The place eluted from silica gel magnetic bead.

Wherein, the cleaning solution can make appropriate choice according to the common knowledge of this field.The one of the cleaning solution A example is：The polyethylene glycol (PEG) of 25mmol/L, the sodium chloride (NaCl) of 3mol/L, 2mmol/L ethyl alcohol.

Wherein, the liquid for being eluted to nucleic acid that the eluent can be known in the art, art technology Personnel can select eluent according to actual conditions.One example of the eluent is：The three hydroxyl first of 10mmol/L Base aminomethane (Tris), the ethylenediamine tetra-acetic acid (EDTA) of 13.7mmol/L.

Therefore, according to a kind of preferred embodiment of the utility model, the nucleic acid purification unit 5 further includes being arranged in core Magnet except piece system, the magnet readily can be introduced or be cut out at any time, so that magnetic bead can basis Demand is fixed and solves fixation.For example, after pyrolysis product enters nucleic acid purification unit 5, need that production will be cracked Object is sufficiently mixed with magnetic bead, so that the nucleic acid in pyrolysis product is fully adsorbed onto on magnetic bead, at this point, magnetic bead needs are in Solve fixed state.And when being cleaned the magnetic bead for being adsorbed with nucleic acid to clean, it needs magnetic bead being fixed so that magnetic Pearl will not because of cleaning solution flushing and as cleaning solution enters next structural unit.

According to the utility model, the nucleic acid amplification unit 4 is nucleic acid (DNA or RNA) molecule progress nucleic acid after purification The place of amplification, the amplification mode can be the cycle temperature control amplification of such as PCR, can also be the constant temperature nucleic acid of such as LAMP Amplification.And when the nucleic acid is RNA molecule, reverse transcription PCR first is carried out to obtain DNA molecular to RNA herein.

Wherein, the nucleic acid amplification occurred in the nucleic acid amplification unit 4 can be reverse transcription PCR, PCR that can also be common To carry out qualitative detection, quantitative fluorescent PCR is can also be to carry out quantitative check.So the region is preferably compared using translucency Prepared by good material, such as polymetylmethacrylate, polycarbonate or polydimethylsiloxane etc..

Wherein, the module that optical detection is carried out to the product after amplification can be provided by external necessary instrument 9.

Wherein, it is provided with the array being made of multiple micro- wells 41 in the nucleic acid amplification unit 4,41 inside of micro- well can be with It is fixed with reverse transcription primer and a variety of different amplimers by the way that point sample instrument point sample is selectable before chip package, this draws For object by being fixed on micro- bottom after particular design, the Primer type in each of micro- well array micro- well 41 can be identical, Can be different, when identical, a type of pathogen is only detected, when different, you can while detecting a variety of different types of Pathogen.Wherein, the size of micro- well 41 can be 0.001-1mm, and shape can be rectangle, round, triangle, diamond shape etc. It is variously-shaped.Array way can be rectangular array, honeycomb type array, circular array etc..

Wherein, when carrying out nucleic acid amplification, required temperature can be provided by external necessary instrument 9.Therefore, described Necessary instrument 9 further includes temperature control module.

According to the utility model, the micro-fluidic chip further includes the waste liquid for collecting waste liquid caused by each step reaction Unit 6, for example, the collection to extra sample to be checked in cell cracking unit 3, is originally quantified, nucleic acid amplification with treating sample Extra purified product is collected in unit 4, to the collection of the lysate after absorption nucleic acid in nucleic acid purification unit 5, to cleaning The collection etc. of liquid and eluent.Preferably, the cell cracking unit 3, nucleic acid amplification unit 4 and nucleic acid purification unit 5 It is respectively arranged with micro-valve door on the micro- access being connected to the waste unit 6, to control the feelings that respective liquid enters waste unit 6 Condition.

Preferably, the connector between the microchannel that each structural unit is communicated with waste unit 6 and waste unit 6 is located at useless The top of liquid unit 6, main function are that the waste liquid after the completion of reacting or extra sample to be checked are transported to waste unit 6 In, simultaneously because the structural relation on position to flow back by microchannel into the waste liquid inside waste unit 6 Part before chip.

Wherein, the shape of the cross section of the microchannel can be rectangle, round, triangle, trapezoidal or other various shapes Shape.The width of microchannel 215 can be 0.001mm-10mm, and depth can be 0.001mm-10mm.

According to the utility model, the waste unit 6 is preferably placed at the end of liquid flow path system in chip, each to collect The waste liquid for walking biochemical reaction prevents waste liquid outflow chip and pollutes external environment.Wherein, the waste unit 6 in structure not It is confined to rectangle shown in Fig. 1, other shapes, such as cylinder, it is conical or other various irregular shapes also the same It is applicable in.Wherein, the volume of the waste unit 6 should be greater than 2 interior reagent volume of all liquid storage units and sample to be checked The sum of volume, to can guarantee that waste unit 6 can effectively collect issuable waste liquid in reaction process.

Wherein, the micro-valve door is preferably provided with piston structure, and opening or closing can be completely by instrument according to predetermined Program is completed.Usually not under the intervention of instrument, micro-valve door is in opening state or part micro-valve door is in the open state, And when needing to be turned off, the piston of micro-valve door is driven by the shaft of stepper motor in instrument, and piston is rotated by a certain angle, So that the fluid path of internal piston is closed so that the transformation of micro-valve door is in off state.It is preferred, therefore, that necessary instrument 9 It further include stepper motor.

As a kind of alternative embodiment, the piston can be vertical slide and non-rotating, in general state The piston of lower micro-valve door is in the open state, and when needing to close, the piston at micro-valve door is pushed by the push-pull rod of instrument, will Piston is pushed into inside microchannels to block microchannel so that the micro-valve door is closed.

As another alternative embodiment, which can also be beaten by the cooperation of iron plate and electromagnetic field Open and close operation places electromagnet, one for example, a small iron plate can be pasted above micro-valve door below micro-valve door As under state, electromagnet no power, iron plate is located above micro-valve door, and micro-valve door is in the open state, when needing to close, passes through Instrument to be powered to electromagnet, and the electromagnetic field that magnet generates can attract the iron plate above micro-valve door, to move iron plate to micro-valve Door lower section so that micro-valve door is closed.

According to the utility model, the sample injection unit 1 is used to receive the sample to be checked of user's injection.The sample to be checked can Think the sample of any required progress PCR amplification, for example, can be but be not limited to, whole blood, serum, blood plasma, urine, saliva, sweat The various body body fluid such as liquid can also be that can carry out the secretion by the secretion of the various histoorgans of body Liquid materials after dilution can also be the cells such as various body tissues, bacterium, virus or other pathogens.

Wherein, the structure of the sample injection unit 1 can be cylindrical, conical (for example, structure shown in Fig. 1), ladder Shape or other various irregular shapes.Its major function is to receive the sample to be checked of user's injection so that sample to be checked is being noted The leakage of sample to be checked will not occur after entering to chip, sample can only be operated according to specific microchannel structure.

As an alternative embodiment, the sample injection unit 1 can be made into capillarity sampling structure, note at this time The sample to be checked entered can wick themselves into the structural unit in downstream, and sample to be checked can be adsorbed on by capillarity In chip, prevents the leakage of sample to be checked and pollute environment.

According to the utility model, the liquid storage unit 2 is at least one, and Fig. 1 shows 5 liquid storage units 2, the liquid storage list The main function of member 2 be the cracking that participate in sample to be checked, purifying, nucleic acid amplification various reaction reagents, for example cell splits It is new to be loaded into this practicality in advance for reagent etc. needed for solution liquid, nucleic acid cleaning solution, DNA/RNA eluents, DNA amplifications or reverse transcription PCR The chip interior of type, when the utility model chip carries out the test of sample to be checked, then will be inside the liquid storage unit 2 by external force Reaction reagent release in sequence.Herein it should be noted that the size and shape of each liquid storage unit 2 of chip interior Shape may be the same or different, and shape is also not limited to circle shown in figure, other shapes such as rectangle, diamond shape, Polygon or irregular shape are equally applicable.The size of each liquid storage unit 2 can be according to the reaction reagent to be pre-installed simultaneously Volume size and change, the volume of liquid storage unit 2 and differ at this time.

As shown in Figure 4 A, the utility model provides a kind of basic structure of liquid storage unit 2, and prepackage reaction reagent is pre- It is first loaded into reagent pouch 21, which is preferably flexible material, which includes but not limited to：Nitrocellulose Film, plastic film or metal aluminum foil, the plastic film can be but be not limited to polyester, polyethylene terephthalate (PET), at least one of makrolon (PC), polypropylene (PP) and polymethyl methacrylate (PMMA).Its common feature It is that can discharge internal prepackage reaction reagent by way of squeezing or needle pierces.It is loaded into reagent pouch in prepackage reaction reagent After in 21, outlet is sealed to form seal 211 by way of thermoplastic envelope.The seal degree of seal 211 herein Than shallower so that when external force pressurizes reagent pouch 21, seal 211 can rupture.Location hole 212 is for assisting reagent capsule Bag 21 is fixed at the liquid storage unit 2 in chip.When needing to discharge 2 internal-response reagent of liquid storage unit, Ke Yitong The mode in Fig. 4 B is crossed, by squeezing the crush-zone 213 in reagent pouch 21, so that seal 211 is ruptured and discharged Liquid can also puncture the bottom section such as needling structure 214 of reagent pouch 21 by pricker by the way of in Fig. 4 C Place so that inside prepackage reaction reagent releases.

According to a kind of preferred embodiment of the utility model, the utility model passes through between micro- access to chip system Pressure or decompression are carried out to control the flowing of each liquid, to enter another structural unit from a structural unit.Cause This, the micro-fluidic chip further includes the ventilation unit 7 being connect with the waste unit 6, for being carried for micro-fluidic chip system For required external pressure, so that internal liquid runs well under the auxiliary of external pressure.Ventilation unit 7 can also be prevented simultaneously Only the waste liquid in waste unit 6 flows out to chip exterior and pollutes external environment.Although ventilation unit 7 as shown in Figure 1 uses W Type microchannel is designed, but other are designed, such as round, arc, other various structures such as Z-type, which can similarly play, ventilates and prevent Only the effect of waste liquid outflow, these designs should all also be treated as being within the protection scope of the utility model.

Preferably, predetermined substance can also be filled inside the ventilation unit 7, the work for playing ventilation but preventing waste liquid from outflowing With, for example the substance can be aerosol, or ventilative but fluid-tight loose cavernous structure substance.

According to a kind of preferred embodiment of the utility model, the micro-fluidic chip of the utility model preferably passes through outside Air pressure needed for 9 offer system of necessary instrument.It is preferred, therefore, that the micro-fluidic chip further includes being connect with the ventilation unit 7 Tool interface system 8, the tool interface system 8 is used to connect necessary instrument 9 outside flow control chip system, and the necessary instrument 9 provides Air pressure the micro-fluidic chip system is provided to by the ventilation unit 7.Wherein, the size of the external pressure can compare Atmospheric pressure is big, can also be smaller than atmospheric pressure, and the size of the air pressure can be easily adjusted according to specific service condition difference.Together When, the tool interface system 8 is preferably air-tightness, i.e. chip, should when the air pressure regulator with necessary instrument 9 is docked Position is air tight.The function can be realized by plastic sealing ring or other assemblies.The air pressure regulator of necessary instrument 9 can To be air driven pump, gas storage vesica, pulsometer, extrusion pump etc..

According to a kind of preferred embodiment of the utility model, as shown in Figure 2, the necessary instrument 9 includes for carrying For promotion pulsometer 91, gas-guide tube 92 and the air-tightness interface 93 being connect with the ventilation unit 7 of air pressure.When chip to be measured After being put into necessary instrument 9, the stepper motor (not shown) of instrument internal can push pulsometer 91 so that air-tightness Interface 93 is clung at the tool interface system 8 of chip to be measured, and it is air-tightness that this, which is close to mode, can be by being connect in the air-tightness Sealing ring or O-ring are set on mouth 93 to complete.Preferably, the air-tightness interface 93 is tubaeform, in the preferred situation Under, air-tightness interface 93 and tool interface system 8 need not be precisely aligned, it is only necessary to which tool interface system 8 is located at the air-tightness interface 93 Inside.It waits after the completion of being close to, the stepper motor being connect with pulsometer 91 is in the lock state, at this time the position of pulsometer 91 It is fixed.Rotation by another the stepper motor (not shown) being connected with 91 internal piston of pulsometer, can push Or extracting piston, to adjust the air pressure inside pulsometer 91, and then manipulate the air pressure of chip interior.It is needing to chip interior When pressurization, is completed by stepper motor to push piston, on the contrary, when needing to depressurize to chip interior, pass through stepper motor It is realized to extract piston.Chip interior fluid path network can drive fluid to shift due to increase or the reduction of air pressure. After the entire testing process of chip to be measured is completed, by the rotation for the stepper motor being connect with pulsometer 91 come so that this is airtight Property interface be detached from chip.

As shown in Figure 5, second aspect according to the present utility model provides a kind of molecule diagnosis micro-fluidic chip system, The micro-fluidic chip system is made of the upper, middle and lower；

Wherein, media layer damage is that molecule as described above diagnoses micro-fluidic chip；

Wherein, superstructure and understructure close the middle layer for covering；It is provided in superstructure and sample introduction Unit 1 connect sample holes, and with the relief hole corresponding to liquid storage unit 2, for providing external impetus to discharge liquid storage list Reaction reagent in member 2.

It is as shown in FIG. 6, it is provided with sample holes (aperture) above superstructure, is used for the sample-adding of sample to be checked.It is described into Sample hole is communicated with the sample injection unit 1 in media layer damage, to ensure that sample to be checked can enter the sample introduction list by sample holes In member 1.In addition, being additionally provided at least one relief hole (macropore) in superstructure, the relief hole corresponds to media layer damage Liquid storage unit 2, the entrance of the push rod for necessary instrument 9, to provide external pressure for liquid storage unit 2, by liquid storage unit The reagent of the advance enclosed storage in 2 inside releases, and therefore, the necessary instrument 9 of the utility model further includes at least one push-and-pull Bar.The relief hole size and shape is determined by the structure size of the liquid storage pouch 21 of liquid storage unit in media layer damage and shape It is fixed, it is preferred that the diameter of the relief hole can be 0.5mm-50mm, and shape can be round, rectangle, polygon, water chestnut Shape or even irregular shape etc. are variously-shaped.

Preferably, the material of the upper, middle and lower is each independently selected from such as dimethyl silicone polymer (PDMS), gathers Methyl methacrylate (PMMA), makrolon (PC), polypropylene (PP), polyethylene terephthalate (PET), plastics are thin Film, elastic emulsion, natural rubber, plastics and silica gel.

Preferably, the thickness of superstructure is 0.5-20mm, more preferably 0.5-10mm, further preferably 0.5mm- 5mm；The thickness of media layer damage is 0.5-50mm；More preferably 2mm-20mm, further preferably 3mm-15mm；Understructure Thickness is 0.5-20mm, more preferably 0.5-10mm, further preferably 0.5mm-5mm.

A kind of preparation method of preferred molecule diagnosis micro-fluidic chip provided by the utility model preferably includes to walk as follows Suddenly：

Step 1) cracks sample to be checked, purify and nucleic acid amplification needed for reaction reagent be preloaded onto the reagent pouch 21 In, and be sealed the seal 212 of the reagent pouch 21 by way of thermoplastic envelope；And it is placed by location hole 211 At the specific liquid storage structure of chip system media layer damage, further by the reagent pouch 21 of each liquid storage by way of thermoplastic envelope It is fixed on chip system media layer damage.

Step 2) places a certain amount of grinding microballon in the first chamber 32 of chip, and in the nucleic acid purification of the chip A certain amount of silica gel magnetic bead is placed in unit 5.

Step 3) fits to understructure below media layer damage as above, and laminating type includes but are not limited to ultrasound The modes such as hot melt, gluing, ultraviolet light solidification, thermoplastic envelope.

Superstructure is fitted to the upper surface of media layer damage as above by step 4), and laminating type includes but are not limited to ultrasound The modes such as hot melt, gluing, ultraviolet light solidification, thermoplastic envelope.

Using a kind of stream of preferred molecule diagnosis micro-fluidic chip system progress molecule diagnosis provided by the utility model Journey is as follows：

Step 1) draws a certain amount of sample to be checked by pipettor, and sample to be checked is passed through the utility model chip system The well of superstructure is added in this chip system, and is entered in sample injection unit 1, then the utility model chip system is put It sets and starts to test in necessary instrument 9.

Step 2) portion in the chip, by internal stepper motor by pulsometer 91, air-tightness interface 93 moves necessary instrument 9 It is fixed after at tool interface system 8 on to chip, then piston is stripped by stepper motor, sample to be checked is inhaled at this time Into the first chamber 32 in cell cracking unit 3, extra sample enters second chamber 33, and under the extracting external application of external force Into waste unit 6, originally quantified to treat sample.

Step 3) closes the micro-valve door of cell cracking unit 3 by necessary instrument 9, and passes through the push-pull rod of necessary instrument 9 Squeezing the lysate pouch in liquid storage unit 2 keeps its internal preloaded with liquid of rupture release, the push-pull rod by necessary instrument 9 continuous Cell cracking unit 3 is squeezed, promotes internal sample to be checked that cell rupture occurs.The micro-valve door of cell cracking unit 3 is opened, and is closed The micro-valve door of closed kernel acid purification unit 5 discharges the cleaning solution in the cleaning solution pouch of liquid storage unit 2, makes it in the same way It rinses product of cell lysis and enters in nucleic acid purification unit 5, fully after cleaning, magnetic is introduced in 5 lower portion of nucleic acid purification unit Silica gel magnetic bead is fixed in iron attraction, opens the micro-valve door of nucleic acid purification unit 5, will cleaning by the extracting effect of necessary instrument 9 Liquid is drawn into waste unit 6, and same mode discharges cleaning solution again, is finally discharged the eluent in liquid storage unit 2, is made The DNA or RNA adsorbed on silica gel magnetic bead is eluted.

Step 4) closes the micro-valve door of cell cracking unit 3 and nucleic acid purification unit 5, and squeezing nucleic acid purification unit 5 makes The DNA solution being eluted inside it enters in nucleic acid amplification unit 5, discharges the nucleic acid amplification agents in liquid storage unit 2, leads to The temperature control for crossing necessary instrument 9 carries out optional reverse transcription PCR and real-time quantitative PCR reaction (or constant temperature LAMP amplifications is anti- It answers).

Step 5) detects fluorescence intensity change in amplification procedure by the light detection module in necessary instrument 9, to Specific amplification curve is obtained, the infection whether sample to be checked occurs certain pathogen is parsed from amplification curve.

The sample volume of molecule diagnosis micro-fluidic chip system detection provided by the utility model is 10-200 μ L.

In the utility model, necessary instrument 9 is small portable apparatus, and necessary instrument 9 is in addition to including pulsometer 91, air guide Further preferably include push-and-pull bar unit, stepper motor module, light detection module, temperature control except pipe 92 and air-tightness interface 93 Module, electromagnet module etc..

The third aspect of the utility model additionally provides molecule diagnosis micro-fluidic chip as described above and/or point Application of the son diagnosis micro-fluidic chip system in nucleic acid amplification.

The utility model will be described in detail by embodiment below.

Embodiment 1

The present embodiment is used to illustrate to detect by using the utility model micro-fluidic chip system the presence of Escherichia coli

The present embodiment using Escherichia coli as the main reason for experiment material is that bacillus coli gene structure understands, and carefully Bacterium culture is simple, and thalline is easy to obtain, and thalline is cheap etc..Certainly equally may be used using experimental method disclosed in the present embodiment For the detection of other pathogens, such as the detection of pathogenic bacteria in septicemia, the detection etc. of respiratory tract infection germ.

The preparation method of cell pyrolysis liquid used in the present embodiment is：Solution is prepared using sterile water, makes its each ingredient Ultimate density be：The dodecyl naphthenic acid sodium (SDS) of 55mmol/L, the ethylenediamine tetra-acetic acid (EDTA) of 25mmol/L, The trishydroxymethylaminomethane (Tris) of 100mmol/L, the sodium chloride (NaCl) of 500mmol/L, is finally adjusted with NaOH solution The pH value of above-mentioned solution is to 8.0, high pressure sterilization 10 minutes.It is cooled to room temperature after the completion of cell pyrolysis liquid sterilizing, then sterile It is encapsulated into environment in liquid storage pouch.

The preparation method of nucleic acid cleaning solution is：Solution is prepared using sterile water, makes the ultimate density of its each ingredient be： The polyethylene glycol (PEG) of 25mmol/L, the sodium chloride (NaCl) of 3mol/L, 2mmol/L ethyl alcohol.Nucleic acid cleaning solution is equal in mixing It is encapsulated into liquid storage pouch in gnotobasis after even.

The preparation method of nucleic acid eluents is：Solution is prepared using sterile water, makes the ultimate density of its each ingredient be： The trishydroxymethylaminomethane (Tris) of 10mmol/L, the ethylenediamine tetra-acetic acid (EDTA) of 13.7mmol/L.Nucleic acid eluents exist It is encapsulated into gnotobasis after mixing in liquid storage pouch.

Quantitative fluorescent PCR reagent is purchased from SYBR Green Mix companies, and main component is：SYBR Green1 dyestuffs, DNTP, Taq archaeal dna polymerase, distilled water.PCR reagent is encapsulated in after being ready to complete in liquid storage pouch 21 in an aseptic environment.

The detection object of the present embodiment is Escherichia coli, and bacterium solution preparation process is：Tryptone 10g is weighed, yeast carries Object 5g, sodium chloride 10g are taken, water is added to be settled to 1L, high pressure steam sterilization 10 minutes is inoculated with exponential phase in Biohazard Safety Equipment Escherichia coli, shaking table culture 16 hours (37 DEG C of shaking table temperature) after inoculation.

The glass microballoon that grinding microballon is diameter 1mm used in the present embodiment, ball shape is hard, spherical preferable, It is added in 2g glass microballoons to the first chamber 32 in the utility model chip.Silica gel magnetic bead is straight used in the present embodiment Diameter is 10 μm or so, and interior nuclear magnetism is superparamagnetism, and shell is covered with macromolecule inert material styrene, when in use first The silica gel magnetic bead is dissolved in aqueous solution, then takes 200 μ L point samples at the nucleic acid purification unit 5 in the chip.

Step 1：The assemble method of chip

1) above-mentioned each reaction reagent is mounted in reagent pouch 21 in advance, and by the reagent pouch by way of thermoplastic envelope 21 seal 212 is sealed (heating time 30s)；And the spy of chip system media layer damage is placed on by location hole 211 Determine at liquid storage structure, the reagent pouch 21 of each liquid storage is further fixed on chip system media layer damage by way of thermoplastic envelope On.

2) upper layer 1 of the utility model chip, middle layer 2 and lower layer 3 are prepared by way of injection molding.First will Each reagent pouch 21 that step 1) obtains is loaded at 2 structure of corresponding liquid storage unit of middle layer 2, then grinding microballon is added It takes 200 μ L to be injected into the nucleic acid purification unit 5 of chip into the first chamber 32 of the chip, then by silica gel magnetic bead, passes through Silica gel magnetic bead is fixed in nucleic acid purification unit 5 by the mode that high temperature is dried to (70 DEG C, 2 hours).Draw needed for quantitative fluorescent PCR Object is by micro- well 41 in point sample instrument point sample to nucleic acid amplification unit 4, and curing 2 hours makes it fix.

Lower layer 3 is fitted in interlayer structure by the cured mode of light-sensitive emulsion (ultraviolet light hardening time 10min) again, Upper layer 1 is fitted in middle layer 2 by same light-sensitive emulsion curing mode (ultraviolet light hardening time 10min), is so prepared Go out complete available chip system.

Step 2：Pattern detection

The pattern detection process of the utility model chip is broadly divided into five steps.

Step 1) draws the Escherichia coli bacteria liquid of 50 μ L by pipettor, and Escherichia coli bacteria liquid is passed through the utility model core The well of piece is added in this chip system, hence into sample injection unit 1, then the utility model chip is placed on mating Start to test in instrument 9.

Step 2) is inside necessary instrument 9, and instrument adjusts the air pressure of chip interior by internal pulsometer 91, by core Sample to be tested inside piece is drawn into the first chamber 32 in cell cracking unit 3, and extra sample is flowed into second chamber In 33, quantifying for sample volume is carried out here, extra sample to be tested is drawn into waste unit 6 by pulsometer.

Step 3) closes the micro-valve door of cell cracking unit 3 by necessary instrument 9, and squeezes lysate pouch and make its rupture The internal preloaded with liquid of release promotes internal sample to be checked that cell occurs broken by the continuously extruded cell cracking unit 3 of push-pull rod It splits.The micro-valve door of cell cracking unit 3 is opened, and closes the micro-valve door of nucleic acid purification chamber 5, is discharged clearly by same mode Cleaning solution in washing lotion pouch makes it rinse product of cell lysis and enters in nucleic acid purification chamber 5, fully after cleaning, in nucleic acid 5 lower portion of purification chamber introduces magnet attraction and fixes silica gel magnetic bead, opens the micro-valve door of nucleic acid purification chamber 5, passes through instrument Extracting effect cleaning solution is drawn into waste unit 6, same mode discharges and discharges cleaning solution again, finally discharges liquid storage In eluent to purification chamber in unit 2, the DNA adsorbed on the silica gel magnetic bead that makes is eluted.

Step 4) closes the micro-valve door of cell cracking unit 3 and nucleic acid purification unit 5, and squeezing nucleic acid purification unit 5 makes it The DNA solution that inside is eluted enters in nucleic acid amplification unit 4, discharges the nucleic acid amplification agents in liquid storage unit 2, passes through The temperature control of necessary instrument 9 carries out fluorescence real-time quantitative PCR reaction, and (its temperature and time is respectively 94 DEG C, 1 minute, then 55 DEG C, 1 minute, then 72 DEG C, 1 minute, carry out at least 20 cycles).

For step 5) in amplification procedure, the SYBR Green fluorescent dyes used in the system can be specifically bound to double-strand On DNA, with the progress of PCR amplification, reagent double center chain DNA is more and more, the SYBR dyestuffs with double-stranded DNA specific binding More and more, obtained fluorescence signal is more and more stronger, and fluorescence intensity is detected by the light detection module in necessary instrument Variation, can obtain specific amplification curve, and the infection that sample to be checked whether there is Escherichia coli is parsed from amplification curve.

Using same testing process, taking sterile waters of the 50 μ L without containing Escherichia coli, as a contrast in addition sample is added to It in one detection chip system, repeats the above steps and is detected, obtain its testing result.

The result shows that this micro-fluidic chip is the positive to the testing result of Escherichia coli solution, and to being free of Escherichia coli Sterile water its testing result be feminine gender, show that the micro-fluidic chip system can be used for carrying out bacterium presence or absence in sample Detection, negative control and positive test show that this detection chip has result accurate, and detection speed is fast (about 50 minutes), can be with It is further used as the molecule diagnostic medicine reference of nucleic acid amplification.

Preferred embodiments of the present invention described in detail above, still, the utility model is not limited to this.At this In the range of the technology design of utility model, a variety of simple variants can be carried out to the technical solution of the utility model, including each Technical characteristic is combined with any other suitable method, these simple variants and combination equally should be considered as the utility model Disclosure of that belongs to the scope of protection of the utility model.

Claims (10)

1. a kind of molecule diagnoses micro-fluidic chip, which is characterized in that the micro-fluidic chip includes：For receive sample to be checked into Sample unit (1), the liquid storage unit (2) for storing reaction reagent, the cell cracked for treating the cell in sample sheet Crack unit (3), the nucleic acid amplification unit (4) for being expanded to nucleic acid；

Wherein, the cell cracking unit (3) includes slit (31) and the first chamber (32) that is separated by the slit (31) With second chamber (33), it is micro- that at least one of the first chamber (32) and the second chamber (33) chamber are configured with grinding Pearl；The first chamber (32) communicates with the sample injection unit (1), the second chamber (33) and at least one liquid storage unit (2) it communicates, for receiving cell pyrolysis liquid.

2. micro-fluidic chip according to claim 1, which is characterized in that the cell cracking unit (3) further includes baffle (34), the baffle (34) is opposite with the slit (31) but does not connect, and the first chamber (32) and second chamber (33) are logical It crosses the baffle (34) and is communicated with the space that the slit (31) defines.

3. micro-fluidic chip according to claim 1 or 2, which is characterized in that the micro-fluidic chip further includes that setting exists Nucleic acid purification unit (5) between the cell cracking unit (3) and nucleic acid amplification unit (4), to enter the core in nucleic acid The pyrolysis product of the cell cracking unit (3) is purified before sour amplification unit (4), the nucleic acid purification unit (5) In configured with capableing of the magnetic bead of specific adsorption nucleic acid.

4. micro-fluidic chip according to claim 1 or 2, which is characterized in that the nucleic acid amplification unit (4) includes by more The array of a micro- well (41) composition is fixed at least one primer pair in micro- well (41).

5. micro-fluidic chip according to claim 1 or 2, which is characterized in that the micro-fluidic chip further includes for receiving Collect the waste unit (6) of waste liquid caused by each step reaction.

6. micro-fluidic chip according to claim 5, which is characterized in that the micro-fluidic chip further includes and the waste liquid list The ventilation unit (7) of first (6) connection, for providing required external pressure for micro-fluidic chip system.

7. micro-fluidic chip according to claim 6, which is characterized in that the micro-fluidic chip further includes single with the ventilation The tool interface system (8) of first (7) connection, the tool interface system (8) are used to connect the necessary instrument (9) outside flow control chip system, institute The air pressure for stating necessary instrument (9) offer is provided to the micro-fluidic chip system by the ventilation unit (7).

8. micro-fluidic chip according to claim 7, which is characterized in that the necessary instrument (9) includes for providing gas Pulsometer (91), gas-guide tube (92) and the air-tightness interface (93) being connect with the ventilation unit (7) of pressure；The air-tightness Interface (93) is tubaeform.

9. micro-fluidic chip according to claim 1 or 2, which is characterized in that the reaction reagent passes through reagent pouch (21) it is loaded into the liquid storage unit (2)；The reagent pouch (21) includes for being fixed to the liquid storage unit (2) On location hole (211), seal (212) and for by the seal (212) open and discharge the reaction reagent Crush-zone (213) or needling structure (214).

10. a kind of molecule diagnoses micro-fluidic chip system, which is characterized in that the micro-fluidic chip system by upper layer, middle level and under Layer composition；

Wherein, media layer damage is that the molecule described in any one of claim 1-9 diagnoses micro-fluidic chip；

Wherein, superstructure and understructure close the middle level for covering；It is provided in superstructure and sample injection unit (1) The sample holes of connection, and with the relief hole corresponding to liquid storage unit (2), for providing external impetus to discharge liquid storage unit (2) reaction reagent in.