CN102296027B - Functional module with replaceable parts for PCR (Polymerase Chain Reaction) amplification - Google Patents
Functional module with replaceable parts for PCR (Polymerase Chain Reaction) amplification Download PDFInfo
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- CN102296027B CN102296027B CN 201110237159 CN201110237159A CN102296027B CN 102296027 B CN102296027 B CN 102296027B CN 201110237159 CN201110237159 CN 201110237159 CN 201110237159 A CN201110237159 A CN 201110237159A CN 102296027 B CN102296027 B CN 102296027B
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Abstract
The invention discloses a functional module with replaceable parts for PCR (Polymerase Chain Reaction) amplification, wherein replaceable microfluidic glass sheets are used as main reaction regions; regions from constant temperature fluid pumped from the outside to hydraulic control constant temperature glass sheets are used as temperature region heat sources; reagent exchange with the outside is carried out through fluid inlet and outlet glass sheets; structural lateral columns of transfer fluid flow passages are used for fixing; the center of the functional module is provided with a center fixed block and heat insulation rubber sheets for filling; the top of the functional module is provided with a fluid control tube for controlling the fluid flow speed; and the fluid control tube can be connected to an outer fluid inlet and outlet control module. The module is convenient to align, match and fix on the aspect of structural design. A spiral flow path can be formed for performing the continuous flow PCR, wherein the circulating times of the spiral flow path can be changed and the fluid in the spiral flow path flows through PCR constant temperature regions sequentially. The functional module has high PCR efficiency, has more answerable conditions, is convenient to disassemble and clean, is easy for large-scale production and integration and can be used under different conditions as required.
Description
Technical field
What the present invention relates to is a kind of functional module of the field-replaceable unit for pcr amplification, belong to biomedical detecting instrument field, specifically a kind of take pcr chip as main body, in constant temperature hydraulic control warm area processed, carry out PCR reaction, can be simultaneously the functional instrument module of integrated or independent use.
Background technology
Polymerase chain reaction (Polymerase Chain Reaction, PCR), also claim acellular clone technology (Free bacteria cloning technique), as molecular biology and an engineered important technology, be a kind of under primer guiding the method for selective amplification DNA and RNA fragment.That round pcr has is special, responsive, productive rate is high, quick, easy, good reproducibility, the easy outstanding advantages such as automatization, can be in small vessels the goal gene that will study or a certain dna fragmentation being increased within a few hours is copied to hundreds of thousands of and even millions of times, for analysis and research with detect and identify.
Pcr amplification needs reaction solution high-temperature denatured, low-temperature annealing, and middle temperature is extended between these three warm areas and is constantly changed, and therefore fast and accurately circulating temperature control is crucial.Need that amount of reagent is large, product must shift the shortcomings such as separation thereby large lifting temperature that there is thermal inertia in traditional round pcr is slow, reaction temperature profile is inhomogeneous, reaction volume is large.Studies show that PCR reaction is very short in sex change and required time of annealing stage in fact, can reduce in a large number the heating and cooling time to accelerate whole reaction process if can overcome thermal inertia in the PCR reaction.
Continuous Flow PCR carries out PCR reaction by making reaction solution flow through different constant temperature warm areas, can overcome the weakness of normal PCR, greatly reduces the reaction times and to the consumption of reagent.This patent further expands and innovates on the basis of micro-fluidic Continuous Flow PCR and capillary type Continuous Flow PCR, be difficult for changing for existing Continuous Flow PCR, also be difficult for modularization and the weakness such as use in batches, external structure is combined with micro-fluidic chip, has stable outside liquid humidity province and unified flow control.One-piece construction is detachably changed, and has the modularization characteristics, is convenient to integrated use and uses separately.
Summary of the invention
The objective of the invention is for the deficiencies in the prior art, a kind of functional module that can pass in and out with external fluid the integrated field-replaceable unit that is used for pcr amplification of control module (controlling the gentle source of the outside pump housing), fluid temperature cycle control module (control constant temperature water tank temperature) is provided.The volution stream that reaction reagent consists of through a plurality of PCR reaction constant temperature warm areas in functional module carries out fast PCR reaction, and can control the number of times of PCR circulating reaction.
The present invention is achieved through the following technical solutions, and the present invention includes: combination functional zone, top, bottom Connection Block, fluid outlet pipe, fluid choke plug, top fluid control tube.Wherein combination functional zone in top are positioned on the top of bottom Connection Block, and combination functional zone, top directly are connected with fluid outlet pipe, fluid choke plug; Combination top, functional zone, top is inserted in fluid control tube bottom, top.
Combination functional zone, described top are from the inside to surface respectively by central fixed block, heat insulation rubber plate, be provided with the removable micro-fluidic sheet glass of fluid channel, be provided with the fluid turnover sheet glass of fluid channel, the hydraulic control constant temperature glass sheet and the structure side column combination that are provided with the fluid channel form, described central fixed block is positioned at combination functional zone, top central authorities, central authorities four right angles of fixed block are fixed wtih the structure lateral column, central authorities are close to by heat insulation rubber plate the fixed block side, and a heat insulation rubber plate in the clamping of per two structure side intercolumniations, a heat insulation rubber plate and a fluid turnover sheet glass in micro-fluidic sheet glass and hydraulic control constant temperature glass sheet or the clamping of structure side intercolumniation.
Described removable micro-fluidic sheet glass, fluid turnover sheet glass, the structure lateral column is the fluid mass component parts, all there is the identical fluid channel of multiple tracks shape component parts inside, above-mentioned fluid mass, form a spiral type stream after the assembling, fluid is by fluid turnover sheet glass turnover stream.
There is the fluid channel described hydraulic control constant temperature glass sheet inside, and the fluid channel is connected with the outside temperature control module with fluid outlet by fluid intake, and replacement fluids constantly flows into the temperature that constant temperature fluid keeps monoblock control constant temperature glass sheet by the external world.
The liquid storage groove is arranged at described fluid turnover sheet glass top, and there are a plurality of porose projections a side, and the liquid storage groove all links to each other with the fluid channel of fluid turnover sheet glass inside with porose projection, and fluid passes in and out side that sheet glass contains porose projection outwardly.
The material of described central fixed block is ABS resin, and heat insulation rubber plate material is the engineering heat resistant rubber.Removable micro-fluidic sheet glass, hydraulic control constant temperature glass sheet, fluid turnover sheet glass main part all is silica glass, there is the fluid channel centre.The porose projection that fluid turnover sheet glass is provided with is the PTEE material.Structure lateral column one-piece construction is ABS resin, and porose projection and inner fluid channel are the PTEE materials.
Described bottom Connection Block is made by ABS resin, and the bottom hollow out has the corresponding screw hole fixing with the structure lateral column of combination functional zone, top on it, bottom Connection Block and combination functional zone, top can be fixed by screw.The porose rectangular preiection corresponding with top combination functional zone hydraulic control constant temperature glass sheet bottom flow turnover rectangular opening arranged, and circular hole that can be by inside and extraneous fluid pump into and pump pipeline and be connected.
Described fluid outlet pipe, fluid choke plug insert in the porose projection on the fluid turnover sheet glass of combination functional zone, top, describedly porosely are raised with N, and then the fluid choked flow is plugged with N-1, and the fluid outlet pipe has 1; Fluid outlet pipe and fluid choke plug position are replaceable, and these two parts determine the final cycle index of fluid spiral type stream together, and outlet pipe is as the outlet of spiral type stream.
Described fluid outlet pipe and described fluid choke plug are made by ABS resin.Both be inserted in the corresponding porose rectangular preiection on the plane of fluid turnover sheet glass of top combination functional zone, its position can be changed according to the actually operating demand.Fluid can flow out by outlet pipe, and the fluid choke plug can stop and answers the hole fluid to flow out.
Fluid control tube top, described top is the pipe that can be connected with extraneous source of the gas or the pump housing, and the fluid that the bottom directly is inserted into combination functional zone, top passes in and out in the liquid storage groove at sheet glass top.There are two screw holes corresponding with the screw hole at the fluid turnover sheet glass top of combination functional zone, top on the fixing crossbeam, top fluid control tube and combination functional zone, top can be fixed by screw.Main part is made by PTEE, has the fixed cross beam of screw hole to be made of metal.
It is as follows to adopt the present invention to carry out PCR reaction working mechanism: (following comprising four blocks of heat insulation rubber plates, three blocks of removable micro-fluidic sheet glass, a fluid turnover sheet glass, the functional module of four structure lateral columns is the example explanation)
In advance reaction reagent is injected into the liquid storage groove at fluid turnover sheet glass top by top fluid control tube, then other one of the pipeline of top fluid control tube is terminated to high-pressure air source or the pump housing, drive the channel flow that reaction reagent enters sheet glass inside by high-pressure air source or the pump housing.By regulating source of the gas or the velocity of flow of pump housing capable of regulating reaction reagent in passage.
Fluid channel in the fluid channel in the fluid turnover sheet glass, three blocks of removable micro-fluidic sheet glass, and the fluid channel part of the structure lateral column between these four blocks of sheet glass has consisted of the fluid environment of reaction reagent jointly.The fluid channel mouth on the both sides, fluid channel in the removable micro-fluidic sheet glass (meeting directly is connected with the porose projection of the structure lateral column of corresponding contact) is in the same level face, identical but the non-touching fluid channel layer of multilayer (design example is 40 layers) shape is arranged, and the fluid between different layers can't directly contact in chip; The fluid channel of structure lateral column also is level, but so that turned to 90 ° on the fluid levels, has connected the fluid on two glass-chips; Fluid channel in the fluid turnover sheet glass is then just in time so that fluid exports outflow from lower one deck after this layer entrance flows into.Be that fluid can pass in and out at fluid and finishing once circulation in the sheet glass and enter lower one deck.
Fluid turnover sheet glass innerly has in the horizontal one and goes out head piece every T layer (T is for changing according to demand) fluid channel except the liquid storage groove at top, goes out head piece and communicates with the porose projection of rectangle on the sheet glass side.The porose projection of rectangle is fluid choke plug beyond the Great Wall generally, does not flow out and continues circulation so that real fluid can not go out head piece from branch road.And reach when needing number of times when PCR circulation, at the porose high spot access of its corresponding rectangle fluid outlet pipe, draw the liquid of finishing reaction cycle.
Every block of removable micro-fluidic sheet glass all is clipped between a heat insulation rubber plate and hydraulic control constant temperature glass sheet.Hydraulic control constant temperature glass sheet internal fluid channels is the serpentine configuration of single continuous bending, and two fluid channel mouths are arranged at the bottom, directly link to each other with the correspondence position of bottom Connection Block.Correspondence position at the bottom Connection Block is connected respectively to going out in head piece and the constant temperature water tank of outside recycle pump by two tubes.Use heating unit and temperature-detecting device to guarantee that the liquid stream temperature in the constant temperature water tank is design temperature.The inflow port of recycle pump is received in the water tank.Open recycle pump during work, make liquid enter a fluid passage mouth of hydraulic control constant temperature glass sheet from constant temperature water tank along pipeline, flow out along channel flow and from another fluid passage mouth after entering chip internal, finally get back to constant temperature water tank.In this flow process, heat exchange makes hydraulic control constant temperature glass sheet remain on the temperature constant state of setting.Under actual PCR Working environment, the temperature of three hydraulic control constant temperature glass sheets is set as respectively 94 ° of C(sex change).60 ° of C(annealing) 72 ° of C(extend).The existence of hydraulic control constant temperature glass sheet so that the micro-fluidic sheet glass that is in contact with it also remains basically identical temperature, when the reagent of inside flows, will obtain the heat transmission of sheet glass in micro-fluidic sheet glass passage.Be that the reagent different micro-fluidic sheet glass of flowing through is actually through different PCR temperature of reaction warm areas, react needed temperature of each stage thereby obtained PCR.The heat insulation rubber plate of another contact surface can reduce the heat loss in the reaction process simultaneously.
Integral construction is so that the design cycle number of times is that n (n is the integral multiple that number of plies T in interval between the head piece of fluid turnover sheet glass) fluid can be mobile in the following order under the external world promotes:
Fluid turnover sheet glass liquid storage groove-〉 fluid turnover sheet glass (1 layer)-〉 structure lateral column A(1 layer)-〉 removable micro-fluidic sheet glass A1,94 ° of C sex change (1 layer)-〉 structure lateral column B(1 layer)-〉 removable micro-fluidic sheet glass B1(1 layer), 60 ° of C annealing-〉 structure lateral column C(1 layers)-〉 removable micro-fluidic sheet glass C1(1 layer) 72 ° of C(extend)-〉 structure lateral column D(1 layer)-〉 fluid turnover sheet glass (finishes once circulation, enter 2 from 1 layer)-〉 structure lateral column A(2 layer)-〉 ...-〉 fluid turnover sheet glass (the n layer is finished n circulation)-〉 corresponding n layer go out head piece-〉 fluid outlet pipe-〉 outside vessel or pipeline.Be that integral body has consisted of a volution stream, have three constant temperature warm areas to carry out the PCR reaction in the stream.
The present invention has following advantage than existing conventional PCR instrument and micro-fluidic chip type PCR:
1. have the advantage of the highly effective reaction of the micro-fluidic PCR reaction of continuous flow pattern, the PCR speed of response is more faster than conventional PCR instrument.
2. utilize constant temperature liquid stream to carry out temperature control, specific heat of water holds high, so the temperature homogeneity of warm area is good, and homoiothermy is than directly by resistive heating with directly dispel the heat far better.
3. module is quick detachable, the convenient cleaning and replacing, can adopt according to the actual requirements different micro-fluidic chip sheet glass to react, the shape of passage, length, with the time that flow velocity has directly determined to carry out each PCR circulation, these can change by changing the micro-fluidic chip sheet glass, even can consider to use the micro-fluidic chip of redesign so that integral body increases new function, and can select the cycle index of W-response.
4. modular design both can be with having cooperated of base and the corresponding pump housing and water tank, and top fluid control tube connects the external pump housing or high-pressure air source is directly used; Also a plurality of modules can be used together.The base unification is placed on the fixed position, then the identical warm area of the base of a plurality of modules can be received on the same water vat and the pump housing, top fluid control tube is received on unified high-pressure air source or the pump housing and is together used, can realize that mass operates simultaneously, both can use separately also and can use together in batches.
5. module can also further directly link to each other base with other equipment that design, and the fluid outlet pipe links to each other with other equipment, does not need outside transfer to enter other equipment and also can realize other functions.Realize integrated.
In a word, the functional module that is used for the field-replaceable unit of pcr amplification is a kind of quick detachable, can separately also can be integrated, can change flexibly according to actual needs, and the invention of convenience and high-efficiency can be made outstanding contribution for stdn and the universalization in this pcr chip field.
Description of drawings
Fig. 1 is the integral shaft mapping of the embodiment of the invention;
Fig. 2 is the axonometric drawing of combination functional zone, the top of the having assembled structure of the embodiment of the invention;
Fig. 3 is the vertical view of combination functional zone, the top of the having assembled structure of the embodiment of the invention;
Fig. 4 is the axonometric drawing that the functional zone structure is made up on the top of the decomposition of the embodiment of the invention;
Fig. 5 is the front view of a kind of removable micro-fluidic sheet glass of the embodiment of the invention;
Fig. 6 is the front view of the another kind of removable micro-fluidic sheet glass of the embodiment of the invention;
Fig. 7 is the local axonometric drawing that amplifies of side of the removable micro-fluidic sheet glass of the embodiment of the invention;
Fig. 8 is the front view of the fluid turnover sheet glass of the embodiment of the invention;
Fig. 9 is the axonometric drawing of the fluid turnover sheet glass of the embodiment of the invention;
Figure 10 is the front view of the hydraulic control constant temperature glass sheet of the embodiment of the invention;
Figure 11 is the axonometric drawing of the structure lateral column of the embodiment of the invention;
Figure 12 is a sectional elevation transversely of the structure lateral column of the embodiment of the invention;
Figure 13 is the axonometric drawing of the central fixed block of the embodiment of the invention;
Figure 14 is the axonometric drawing of the heat insulation rubber plate of the embodiment of the invention;
Figure 15 is the axonometric drawing in front of the bottom Connection Block of the embodiment of the invention;
Figure 16 is the axonometric drawing at the back side of the bottom Connection Block of the embodiment of the invention;
Figure 17 is the fluid outlet pipe of the embodiment of the invention, the axonometric drawing at the back side of fluid choke plug;
Figure 18 is the axonometric drawing of the top fluid control tube of the embodiment of the invention.
Among the figure: combination functional zone, 1-top, 2-bottom Connection Block, 3-fluid outlet pipe, 4-fluid choke plug, 5-top fluid control tube, 6-central fixed block, 7-heat insulation rubber plate, 8-removable micro-fluidic sheet glass, 9-fluid turnover sheet glass, 10-hydraulic control constant temperature glass sheet, 11-structure lateral column.
Embodiment
Below in conjunction with accompanying drawing embodiments of the invention are elaborated.Following examples carry out setting forth principle of work with the PCR reaction of 25 circulations as sample, but protection scope of the present invention is not limited to following embodiment.
As shown in Figure 1, present embodiment comprises: combination functional zone 1, top, bottom Connection Block 2, fluid outlet pipe 3, fluid choke plug 4 and top fluid control tube 5.Wherein: combination functional zone 1, top are arranged on the top of bottom Connection Block 2, and both just in time can block mutually surely on the structure design; Fluid outlet pipe 3 is in the porose projection on the side (fluid turnover sheet glass 9 consists of) of combination functional zone 1, fluid choke plug 4 insertion top.Among the embodiment, the porose of correspondence is raised with 8.And fluid choke plug 4 has 7, and fluid outlet pipe 3 has 1, and their position can be moved practical situation according to module and change at any time; In the liquid storage groove of combination functional zone 1, insertion top, fluid control tube 5 bottoms, top (fluid turnover sheet glass 9 tops).
Such as Fig. 2, Fig. 3, shown in Figure 4, combination functional zone 1, described top are from the inside to surface respectively by central fixed block 6, heat insulation rubber plate 7, removable micro-fluidic sheet glass 8, fluid turnover sheet glass 9, hydraulic control constant temperature glass sheet 10, structure lateral column 11 combines, and is fixed by each structural screw hole.Central authorities' fixed block 6 is positioned at combination functional zone, top 1 central authorities, central authorities 6 four right angles of fixed block are fixed wtih structure lateral column 11,6 four faces of central authorities' fixed block are close to by four blocks of heat insulation rubber plates 7, have 11 clampings of two structure lateral columns a heat insulation rubber plate 7 and a fluid turnover sheet glass 9, a heat insulation rubber plate 7, a micro-fluidic sheet glass 8 and a hydraulic control constant temperature glass sheet 10 in 11 clampings of other per two structure lateral columns.In the present embodiment, heat insulation rubber plate 7 has 4, by indicating with 7A, 7B, 7C, 7D counterclockwise; Removable micro-fluidic sheet glass 8 has 3, indicates with 8A, 8B, 8C counterclockwise; Structure lateral column 11 has 4, presses counterclockwise 11A, 11B, 11C, 11D sign.Combination functional zone 1, top are the main functionality parts for the functional module of the field-replaceable unit of pcr amplification.
As shown in Figure 2, behind all component combinations of above-mentioned combination functional zone 1, top, its top is all in a horizontal plane; Heat insulation rubber plate 7, removable micro-fluidic sheet glass 8, fluid turnover sheet glass 9, also all on the same level face, this is highly identical in the bottom of hydraulic control constant temperature glass sheet 10; The bottom of central authorities' fixed block 6 and structure lateral column 11 is in a horizontal plane, and both are highly identical.The height of the aspect ratio of both (central fixed block 6 and structure lateral column 11) front four (heat insulation rubber plate 7, removable micro-fluidic sheet glass 8, fluid turnover sheet glass 9, hydraulic control constant temperature glass sheets 10) wants high afterwards.Because the top alignment, front four bottom forms four depressions than rear both depressions according to block position, and cup depth is corresponding with corresponding difference of altitude.This difference of altitude also just in time is the height (visible Figure 15) of four boss of bottom Connection Block 2
Described bottom Connection Block 2 is made by ABS resin, and such as Figure 15, shown in Figure 16, the bottom hollow out is because the screw hole of four structure lateral column 11 correspondences of combination functional zone, top can make up functional zone with bottom Connection Block and top by screw and fix.Six the porose rectangular preiections (Figure 15) corresponding with the bottom flow turnover rectangular opening of top combination functional zone three hydraulic control constant temperature glass sheet 10 are arranged, and six circular holes at its corresponding back side and extraneous fluid pump into and pump pipeline be connected (Figure 16).Bottom Connection Block 2 Main Functions are to connect outside constant temperature fluid, support fixedly total, and make things convenient for and being connected and fixed of other peripheral equipments.
Described fluid outlet pipe 3 and described fluid choke plug 4 are made by ABS resin, and its structure as shown in figure 17.Both be inserted in the corresponding porose rectangular preiection on the side of fluid turnover sheet glass 9 of top combination functional zone (Fig. 9), its position can be changed according to the actually operating demand.Fluid choke plug 4 is not so that the fluid in the corresponding porose rectangular preiection can flow out, and 3 of fluid outlet pipes can make the fluid in the corresponding porose rectangular preiection be flowed out.Under the normal operation, the reagent that fluid outlet pipe 3 is derived is collected, namely obtained through the finished product of reaction of designated period PCR circulation.
Described central fixed block 6 is a rectangular parallelepiped as shown in figure 13, and end face and bottom surface are squares.Material is ABS resin.When it assembles the described bottom of complete insertion Connection Block 2 in above-mentioned combination functional zone, top 1, just can be stuck in bottom 2 four protruding blocks of Connection Block and surround in the central zone that forms (Figure 15).The major function of central authorities' fixed block 6 has been central supporting effects of combination functional zone 1, whole top.
Described heat insulation rubber plate 7 structures are the structure of a simple shape as shown in figure 14, make with heat resistant rubber with engineering, the notch cuttype structure on its both sides be for the notch cuttype structural correspondence of structure lateral column 11, convenient location is steady with card during assembling.The major function of heat insulation rubber plate 7 is that it is too large that temperature is unlikely to be subjected to extraneous convection current and heat exchange to affect so that the removable micro-fluidic sheet glass 8 temperature homoiothermy that are in direct contact with it keep better.Another function is as the fixing buffering of structure.
Described removable micro-fluidic sheet glass 8 is made by silica glass.Global shape is the rectangle thin slice, such as Fig. 5, shown in Figure 6.There is the fluid channel inside, the right and left is run through in the fluid channel, the place, gangway of the right and left also has custom-designed than the larger a little porthole (Fig. 7) of inner other passages, can with the side of the micro-fluidic sheet glass 8 of structure lateral column 11 clampings on porose projection corresponding (seeing Figure 11) do not reveal so that both fluid channels can engage.The fluid channel is multiple tracks, and is spaced apart with each other, can not directly be connected in sheet glass inside.Be 40 roads in the present embodiment, mean and to carry out maximum 40 PCR loop cycles.(port number of the fluid turnover sheet glass 9 of back is number herewith).Fig. 5 is two kinds of removable micro-fluidic sheet glass 11 that the internal fluid channels part is different from Fig. 6.The use of can corresponding when reagent react, need in the different time situation of different warm areas, arranging in pairs or groups.Embody the characteristics that removable micro-fluidic sheet glass 11 can be changed as required.Micro-fluidic sheet glass 8 is reagent react and the home court institute of flowing.
Described fluid turnover sheet glass 9, structure such as Fig. 8, shown in Figure 9.Be by the center similar to micro-fluidic sheet glass 8 being bonded of the silica glass chip of fluid channel and front outer shroud one circle silica glass pad (be used for interstitital texture lateral column 11 respective clamp positions) to be arranged, the spacer thickness just in time thickness with hydraulic control constant temperature glass sheet 10 is identical.Sagging liquid storage groove is arranged at the top, and the aperture of groove central authorities is the fluid channel of direct fluid turnover sheet glass 9 inside.As shown in Figure 9, similar with micro-fluidic sheet glass 8, the right and left also has the porthole, but the fluid channel of fluid turnover sheet glass 9 inside is asymmetric, when the liquid that the right-side channels mouth flows into flowed out from porthole, the left side, the outlet position was than the entrance location porthole that descended.This design is so that after overall package is good, and reagent will be realized lane change in fluid passes in and out the fluid channel of sheet glass 9, namely finish the passage that a circulation enters next circulation.Fluid turnover sheet glass 9 inside can transversely have one and go out head piece every T layer (T is for changing according to demand, and this embodiment is T=5) fluid channel, go out head piece and communicate with the porose projection of rectangle on the sheet glass side.The porose projection of rectangle is made with the PTEE material.The porose projection outside of rectangle is fluid choke plug 4 beyond the Great Wall generally, stops up this and goes out the head piece branch road, makes fluid continue to circulate in the fluid channel.In order to access needed PCR cycle index so that whole system is reacted, the porose projection of the rectangle of corresponding cycle index will connect fluid outlet pipe 3, and the liquid of finishing reaction cycle is flowed out from here.This embodiment fluid outlet pipe 3 is connected on the 5th rectangle free high spot, and when namely carrying out the PCR reaction by embodiment, reagent will flow out from fluid outlet pipe 3 after having reacted 5*5=25 PCR circulation, finish reaction.
Described hydraulic control constant temperature glass sheet 10 is made by silica glass.Global shape is the rectangle thin slice, as shown in figure 10.There is the fluid channel inside, and the fluid channel forms a serpentine path, and there are two openings the bottom surface, as fluid inlet and outlet.It can correspondingly with the porose rectangular preiection on the bottom Connection Block 2 link to each other, the pump housing that the conduit that final two fluid inlet and outlets are introduced by bottom Connection Block 2 respectively accesses extraneous constant temperature water tank and follows constant temperature water tank.The thermostat(t)ed water that the outside pump housing pumps into will be so that hydraulic control constant temperature glass sheet 10 keeps constant temperature.Hydraulic control constant temperature glass sheet 10 Main Functions are exactly by thermal contact conductance, keep the directly temperature of the removable micro-fluidic sheet glass 8 of contact so that the PCR reaction obtain need to temperature.
Described structure lateral column 11, its structure such as Figure 11, shown in Figure 12.Wherein the structure on the both direction (being mutually 90 ° of C) is used for the heat insulation rubber plate 7 on the both direction, removable micro-fluidic sheet glass 8, and fluid turnover sheet glass 9, adding of hydraulic control constant temperature glass sheet 10 held.As shown in figure 11, porose projection on the medial surface has the fluid channel (Figure 12) that two porose projections are connected mutually in the horizontal plane in the porose projection.Porthole corresponding to removable micro-fluidic sheet glass 8 or hydraulic control constant temperature glass sheet 10 sides.Can when assembling, the fluid channel well be sealed, so that the fluid channel of the sheet glass that structure lateral column 11 two ends are followed is connected.Screw hole on the side be used for add the sheet glass of holding fix, the screw hole of bottom (Figure 11) is used for fixing with the corresponding screw hole of bottom Connection Block 2.Structure lateral column 11 one-piece constructions are made with ABS resin, but internal fluid channels and porose projection are to obtain with filling in structure lateral column 11 after the PTEE materials processing.
Such as Fig. 2, Fig. 3, shown in Figure 4, the assembling mode of combination functional zone 1, described top is as follows: all structural top are on the same level face.Central authorities' fixed block 6 is positioned at central authorities, and the right angle bayonet socket of four structure lateral columns 11 is fitted fully with four right angles of central fixed block 6 respectively.Four heat insulation rubber plate 7(Figure 14), a side of central fixed block 6 is close in the side that each will be less, and the both sides of each heat insulation rubber plate 7 are steady by the counter structure card of structure lateral column 11.This moment 3 blocks of removable micro-fluidic sheet glass 8 are injected respectively the one side by heat insulation rubber plate 7 of the slot of per two structure lateral columns 11 formation.Fluid is passed in and out in the slot that two structure lateral columns 11 that sheet glass 9 do not insert micro-fluidic sheet glass 8 form, contain the side of porose projection outwardly.Then 3 hydraulic control constant temperature glass sheets 10 are inserted into respectively the slot on 3 removable micro-fluidic sheet glass 8 next doors.Finish structure such as Fig. 2 of combination is shown in Figure 3.
Removable micro-fluidic sheet glass 8, fluid turnover sheet glass 9, hydraulic control constant temperature glass sheet 10 has screw hole on the side of structure lateral column 11 these four parts.Screw hole has corresponding relation, after assembling by above-mentioned assembling mode, from the screw hole of the outside surface of structure lateral column 11 screw is torqued into, the removable micro-fluidic sheet glass 8 that structure lateral column 11 is folded with it, hydraulic control constant temperature glass sheet 10 or the structure lateral column 11 fluids turnover sheet glass 9 folded with it can be completely fixed non-loosening.Such as Fig. 1, shown in Figure 11, screw hole is arranged at structure lateral column 11 bottoms, and four structure lateral columns 11 have 4 screw holes, and is corresponding with four screw holes on the bottom Connection Block 2.Top is made up after functional zone 1 insert bottom Connection Block 2, from the screw column (seeing Figure 16) of bottom Connection Block 2 back screw is torqued into, functional zone 1 are made up on top namely be completely fixed non-loosening with bottom Connection Block 2.Top fluid control tube 5 has two screw holes, and its two screw holes with fluid turnover sheet glass 9 tops in section combination function district 1 are corresponding.Fluid control tube 5 bottoms, top are inserted combination functional zone 1, top the liquid storage groove after, two screws of correspondence are torqued into.Such as Fig. 1, shown in Figure 180, combination functional zone 1, top namely are completely fixed with top fluid control tube 5.
After present embodiment is finished and is all assembled, as shown in Figure 1.Then the circular hole of bottom Connection Block 2 corresponding hydraulic control constant temperature glass sheets 10 is plugged pipeline, be connected to the outside pump housing and constant temperature water tank.So that the steady temperature of hydraulic control constant temperature glass sheet 10A, 10B, 10C is set as respectively 94 ° of C(sex change).60 ° of C(annealing) 72 ° of C(extend).Inject the liquid storage groove by top fluid control tube 5 with reaction solution this moment, and top fluid control tube 5 is received outside source of the gas or the pump housing.External air source or the pump housing will so that the reaction solution of liquid storage groove the inside enter by 4 blocks of sheet glass (9,8A, 8B, 8C) and four structure lateral column 11(11A, 11B, 11C, 11D) the liquid that forms of internal fluid channels flow circulation passage.According to 9-〉11A-〉8A-11B-〉8B-〉11C-8C-11D-9-... sequential loop flow, then can pass in and out sheet glass 9 at fluid and enter next circulation, wholely consist of a volution stream.Because removable micro-fluidic sheet glass 8A, 8B, 8C respectively with hydraulic control constant temperature glass sheet 10A, 10B, 10C directly fits, also can the flow through warm area of corresponding hydraulic control constant temperature glass sheet of the reaction solution of these three sheet glass internal fluid channels of therefore flowing through in order.Be that fluid is at every turn by 94 ° of C(sex change).60 ° of C(annealing) 72 ° of C(extensions) sequential flowing of warm area is finished the PCR reaction cycle one time.And whenever carrying out 5 circulations, fluid can be run into a branch road at fluid turnover sheet glass 9 and go out head piece and lead to the porose projection of rectangle in fluid turnover sheet glass 9 outsides.Protrudingly all clogged by fluid choke plug 4 because front 4 rectangles are porose, thus branch road to go out head piece inoperative.What the branch road of the 25th circulation went out the outside connection of head piece (corresponding the 5th porose projection of rectangle) is fluid outlet pipe 3, because branch road is much smaller than the pressure of the internal fluid channels that continues to go ahead, fluid will flow out from fluid outlet pipe 3 after 25 PCR circulations, finishes the PCR reaction process of whole 25 circulations.
To sum up, the present invention proposes a kind of functional module of the field-replaceable unit for pcr amplification, the zone take removable micro-fluidic sheet glass as principal reaction, the constant temperature fluid that pumps into the external world arrives hydraulic control constant temperature glass sheet as each warm area thermal source, carry out the reagent exchange by fluid turnover sheet glass and the external world, take the structure lateral column of the fluid channel of four transfers as fixing, the center is useful on central fixed block and the heat insulation rubber plate of filling, the fluid control tube of control flow stream velocity is arranged at the top, can receive external fluid turnover control module (controlling the gentle source of the outside pump housing).The bottom is the bottom Connection Block that is connected to fluid temperature cycle control module (the outside pump housing or source of the gas) and fixed sturcture.Conveniently aim at and be fixed in the modular structure design, with fixing whole each parts of corresponding screw hole.Can form a cycle index can change, and flow through the successively volution stream of three PCR reaction constant temperature warm areas of fluid is to carry out continuously streamed PCR reaction.Be that a PCR reaction efficiency is high, it is many to tackle situation, for convenience detach and cleaning, and it is integrated to be easy to scale operation, the functional module that can use under different situations as required.
Above embodiment shows in detail and has described ultimate principle of the present invention, principal character and advantage of the present invention.The technician of the industry should understand, and the present invention is not restricted to the described embodiments, and the present invention also has various changes and modifications without departing from the spirit and scope of the present invention, and these changes and improvements all fall in the claimed scope of the invention.The claimed scope of the present invention is defined by appending claims and equivalent thereof.
Claims (8)
1. a functional module that is used for the field-replaceable unit of pcr amplification is characterized in that: comprise combination functional zone (1), top, bottom Connection Block (2), fluid outlet pipe (3), fluid choke plug (4) and top fluid control tube (5); Wherein: combination functional zone, top (1) are positioned on the top of bottom Connection Block (2), and combination functional zone, top (1) directly are connected with fluid outlet pipe (3), fluid choke plug (4); Combination top, functional zone (1), top is inserted in fluid control tube bottom, top;
Described combination functional zone, top (1) are from the inside to surface respectively by central fixed block (6), heat insulation rubber plate (7), be provided with the removable micro-fluidic sheet glass (8) of fluid channel, be provided with the fluid turnover sheet glass (9) of fluid channel, the hydraulic control constant temperature glass sheet (10) and the structure lateral column (11) that are provided with the fluid channel combine; Described central fixed block (6) is positioned at combination functional zone, top central authorities, and central fixed block (6) corner is fixed wtih structure lateral column (11), and heat insulation rubber plate (7) is close in central fixed block (6) side; Clamping between a heat insulation rubber plate (7), a micro-fluidic sheet glass (8) and a hydraulic control constant temperature glass sheet (10) or two structure lateral columns (11) clamping and a heat insulation rubber plate (7) and a fluid turnover sheet glass (9) between per two structure lateral columns (11);
Described central fixed block (6), heat insulation rubber plate (7), removable micro-fluidic sheet glass (8), fluid turnover sheet glass (9), hydraulic control constant temperature glass sheet (10) and structure lateral column (11), the top of these parts is on the same level face, the bayonet socket of structure lateral column (11) is fitted fully with the corner of central fixed block (6) respectively, a side of central fixed block (6) is close in the side that each piece all will be less in the heat insulation rubber plate (7), and the both sides of each heat insulation rubber plate (7) are blocked by the counter structure of structure lateral column (11); Removable micro-fluidic sheet glass (8) is inserted in respectively the one side by heat insulation rubber plate (7) of the slot of per two structure lateral columns (11) formation; Fluid turnover sheet glass (9) does not insert in the slot of two structure lateral columns (11) formation of micro-fluidic sheet glass (8); Hydraulic control constant temperature glass sheet (10) is inserted into respectively the slot on removable micro-fluidic sheet glass (8) next door;
Described removable micro-fluidic sheet glass (8), all there is the identical fluid channel of multiple tracks shape fluid turnover sheet glass (9) and structure lateral column (11) inside, form a spiral type stream after the assembling, and fluid passes in and out sheet glass (9) turnover stream by stream.
2. the functional module of the field-replaceable unit for pcr amplification according to claim 1, it is characterized in that: there is the fluid channel described hydraulic control constant temperature glass sheet (10) inside, the fluid channel is connected with the outside temperature control module with fluid outlet by fluid intake, replacement fluids constantly flows into the temperature that constant temperature fluid keeps monoblock control constant temperature glass sheet (10) by the external world.
3. the functional module of the field-replaceable unit for pcr amplification according to claim 1, it is characterized in that: described hydraulic control constant temperature glass sheet (10) internal fluid channels forms a serpentine path, two fluid channel mouths are arranged at the bottom, directly link to each other with the correspondence position of bottom Connection Block (2); The fluid channel mouth on the both sides, fluid channel in the removable micro-fluidic sheet glass (8) directly is connected with the porose projection of the structure lateral column (11) of corresponding contact, identical but the non-touching fluid channel layer of multilayer shape is arranged, and the fluid of each interlayer can't directly contact in chip; The fluid channel of structure lateral column (11) is level, connects the fluid of same layer on two glass-chips; Fluid channel in the fluid turnover sheet glass (9) is then just in time so that fluid exports outflow from lower one deck after this layer entrance flows into.
4. the functional module of the field-replaceable unit for pcr amplification according to claim 1, it is characterized in that: the liquid storage groove is arranged at described fluid turnover sheet glass (9) top, there are a plurality of porose projections a side, liquid storage groove and porose projection all link to each other with the inner fluid channel of fluid turnover sheet glass (9), and fluid passes in and out side that sheet glass (9) contains porose projection outwardly.
5. the functional module of the field-replaceable unit for pcr amplification according to claim 1, it is characterized in that: described fluid outlet pipe (3), in the porose projection on the fluid turnover sheet glass (9) of fluid choke plug (4) insertion combination functional zone, top (1), the described porose N of being raised with, then fluid choke plug (4) has N-1, and fluid outlet pipe (3) has 1; Fluid outlet pipe (3) and fluid choke plug (4) position are replaceable, and these two parts determine the final cycle index of fluid spiral type stream together, and fluid outlet pipe (3) is as the outlet of spiral type stream.
6. the functional module of the field-replaceable unit for pcr amplification according to claim 1, it is characterized in that: described top fluid control tube (5) one ends connect the external control module, the other end can connect the stream that internal flow zone, top combination functional zone (1) component parts consists of, and makes up velocity of flow in the internal flow path of functional zone (1) on top by external air source or pump housing control fluid.
7. the functional module of the field-replaceable unit for pcr amplification according to claim 1, it is characterized in that: described top fluid control tube (5) top is the flexible pipe that can be connected with extraneous source of the gas or the pump housing, the fluid that the bottom directly is inserted into combination functional zone, top (1) passes in and out in the liquid storage groove at sheet glass (9) top, and the liquid storage groove at fluid turnover sheet glass (9) top is as the entrance of volution stream.
8. the functional module of the field-replaceable unit for pcr amplification according to claim 1, it is characterized in that: the PCR reaction reagent flowed into from fluid passes in and out the liquid storage groove at sheet glass (9) top when this functional module was used, spiral type stream through three constant temperature warm areas, finish the PCR reaction of repeatedly circulation, finally derive from outlet pipe (3).
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| Application Number | Priority Date | Filing Date | Title |
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| CN 201110237159 CN102296027B (en) | 2011-08-18 | 2011-08-18 | Functional module with replaceable parts for PCR (Polymerase Chain Reaction) amplification |
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| CN 201110237159 CN102296027B (en) | 2011-08-18 | 2011-08-18 | Functional module with replaceable parts for PCR (Polymerase Chain Reaction) amplification |
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| CN109022263A (en) * | 2017-06-08 | 2018-12-18 | 昆泰锐（武汉）生物技术有限责任公司 | A kind of tubular type temperature regulating device and the reaction instrument comprising the tubular type temperature regulating device |
| CN107460109A (en) * | 2017-07-23 | 2017-12-12 | 新疆昆泰锐生物技术有限公司 | A kind of reaction system for bacterium colony PCR is prepared and sampling device and PCR instrument |
| CN107446811A (en) * | 2017-07-25 | 2017-12-08 | 新疆昆泰锐生物技术有限公司 | A kind of reaction instrument for PCR tubular type temperature regulating device and comprising the device |
| CN107446812A (en) * | 2017-07-25 | 2017-12-08 | 新疆昆泰锐生物技术有限公司 | A kind of reaction instrument for PCR four-temperature region tubular type temperature regulating device and comprising the device |
| CN107703320B (en) * | 2017-10-27 | 2023-11-24 | 大连量子流体控制技术有限公司 | Fully-integrated multi-channel multifunctional microfluidic analysis experiment system |
| CN109030437B (en) * | 2018-07-04 | 2020-12-11 | 中山出入境检验检疫局检验检疫技术中心 | Detection device and detection method for aflatoxin |
| CN110261310A (en) * | 2019-07-10 | 2019-09-20 | 深圳金迈隆电子技术有限公司 | A kind of on piece laboratory testing device |
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| CN201130152Y (en) * | 2007-12-21 | 2008-10-08 | 北京工业大学 | Apparatus for measuring and controlling speed of microfluid fluorescence in fluorescence PCR micro-current-control chip micro-channel |
| KR101221872B1 (en) * | 2009-04-16 | 2013-01-15 | 한국전자통신연구원 | Apparatus for polymerase chain reaction |
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