CN102296027A - 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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- CN102296027A CN102296027A CN2011102371591A CN201110237159A CN102296027A CN 102296027 A CN102296027 A CN 102296027A CN 2011102371591 A CN2011102371591 A CN 2011102371591A CN 201110237159 A CN201110237159 A CN 201110237159A CN 102296027 A CN102296027 A CN 102296027A
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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 that is used for the field-replaceable unit of pcr amplification, belong to biomedical detecting instrument field, specifically be a kind of, in constant temperature hydraulic control system warm area, carry out PCR reaction based on pcr chip, can be integrated simultaneously or the functional instrument module of independent use.
Background technology
Polymerase chain reaction (Polymerase Chain Reaction, PCR), also claim acellular clone technology (Free bacteria cloning technique), as a molecular biology and an engineered important technology, be a kind of under primer guiding selective amplification DNA and the segmental method of RNA.That round pcr has is special, responsive, productive rate is high, quick, easy, good reproducibility, easy outstanding advantage such as automatization, can be in small vessels the goal gene that will study or a certain dna fragmentation being increased in 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 circulating temperature control fast and accurately is crucial.Need that amount of reagent is big, product must shift shortcomings such as separation thereby big lifting temperature that there is thermal inertia in traditional round pcr is slow, reaction temperature profile is inhomogeneous, reaction volume is big.Studies show that PCR reaction is very short in sex change and required time of annealing stage in fact, can reduce the heating and cooling time in a large number to quicken entire reaction course 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 conventional P CR, significantly 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 at existing Continuous Flow PCR, also be difficult for modularization and weakness such as use in batches, external structure is combined with micro-fluidic chip, have stable external fluid temperature district 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 at the deficiencies in the prior art, a kind of functional module that can pass in and out 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) with external fluid is provided.The volution stream that reaction reagent constitutes through a plurality of PCR reaction constant temperature warm areas in functional module carries out PCR reaction fast, 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 sheet glass 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, the structure lateral column is being fixed in four right angles of fixed block by central authorities, 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 sheet glass 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, the assembling back forms a spiral type stream, and fluid is by fluid turnover sheet glass turnover stream.
There is the fluid channel described hydraulic control constant temperature sheet glass 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 sheet glass by the external world.
The liquid storage groove is arranged at described fluid turnover sheet glass top, and there are a plurality of porose projectioies 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 an ABS resin, and heat insulation rubber plate material is the engineering heat resistant rubber.Removable micro-fluidic sheet glass, hydraulic control constant temperature sheet glass, fluid turnover sheet glass main part all is a 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 an 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 of structure lateral column fixed that makes up functional zone with top on it, bottom Connection Block and combination functional zone, top can be fixed by screw.Have and the corresponding porose rectangular preiection of top combination functional zone hydraulic control constant temperature sheet glass bottom flow turnover rectangular opening, circular hole that can be by inside and extraneous fluid pump into and pump pipeline and be connected.
In the porose projection on the fluid turnover sheet glass of described fluid outlet pipe, combination functional zone, fluid choke plug insertion top, the described porose N that is raised with is individual, 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.The both is inserted in the corresponding porose rectangular preiection on the plane of fluid turnover sheet glass of top combination functional zone, and 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 the 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 on the fixed crossbeam, top fluid control tube and combination functional zone, top can be fixed by screw with the screw hole at the fluid turnover sheet glass top of combination functional zone, top.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 the high-pressure air source or the pump housing, drive the channel flow that reaction reagent enters sheet glass inside by the high-pressure air source or the pump housing.Can adjust the velocity of flow of reaction reagent in passage by regulating the source of the gas or the pump housing.
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 constituted 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 same horizontal plane, 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 a level, but make and turned to 90 ° on the fluid levels, has connected the fluid on two glass-chips; Flow out from following one deck outlet after then just in time making fluid flow into from this layer inlet fluid channel in the fluid turnover sheet glass.Be that fluid can pass in and out at fluid and finishing once circulation in the sheet glass and enter one deck down.
Fluid turnover sheet glass innerly has one in the horizontal 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, makes real fluid can not go out head piece from branch road and flows out and continue circulation.And reach when needing number of times when PCR circulation, insert the fluid outlet pipe at the porose high spot of its pairing rectangle, 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 sheet glass.Hydraulic control constant temperature sheet glass internal fluid channels is the serpentine configuration of single continuous bending, and two fluid channel mouths are arranged at the bottom, directly links 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 sheet glass 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 sheet glass remain on the temperature constant state of setting.Under actual PCR Working environment, the temperature of three blocks of hydraulic control constant temperature sheet glass is set at 94 ° of C(sex change respectively).60 ° of C(annealing) 72 ° of C(extend).The existence of hydraulic control constant temperature sheet glass makes the micro-fluidic sheet glass that is in contact with it also remain the temperature of basically identical, when the reagent of inside flows in micro-fluidic sheet glass passage, will obtain the heat transmission of sheet glass.Be that the reagent different micro-fluidic sheet glass of flowing through is actually and has passed through different PCR temperature of reaction warm areas, react needed temperature of each stage thereby obtained PCR.The heat that the heat insulation rubber plate of another contact surface of while can reduce in the reaction process scatters and disappears.
Integral construction makes that the design cycle number of times is that n (n is the integral multiple that interval number of plies T between the head piece of fluid turnover sheet glass) fluid can flow 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 constituted 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 different micro-fluidic chip sheet glass to react according to the actual requirements, the shape of passage, length has directly determined to carry out each PCR round-robin time with flow velocity, and these can change by changing the micro-fluidic chip sheet glass, even can consider to make integral body increase new function, and can select the cycle index of W-response with the micro-fluidic chip of redesign.
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, the identical warm area of the base of a plurality of modules can be received on the same water vat and the pump housing then, top fluid control tube is received on the 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 convenient invention efficiently can be made outstanding contribution for the 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 having assembled the structure of the embodiment of the invention;
Fig. 3 is the vertical view of combination functional zone, the top of having assembled the 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 sheet glass 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 sheet glass, 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 25 round-robin PCR reactions 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 constitutes) 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).
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 sheet glass 10, structure lateral column 11 combines, and fixes by each structural screw hole.Central authorities' fixed block 6 is positioned at combination functional zone, top 1 central authorities, structure lateral column 11 is being fixed in 6 four right angles of fixed block by central authorities, 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 fluid turnover sheet glass 9, a heat insulation rubber plate 7, a micro-fluidic sheet glass 8 and a hydraulic control constant temperature sheet glass 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 11A, 11B, 11C, 11D sign counterclockwise.Combination functional zone 1 in top are the main functionality parts of functional module that is used for the field-replaceable unit of pcr amplification.
As shown in Figure 2, after all parts 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 same horizontal plane, this is highly identical in the bottom of hydraulic control constant temperature sheet glass 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 back both (central fixed block 6 and structure lateral column 11) preceding four (heat insulation rubber plate 7, removable micro-fluidic sheet glass 8, fluid turnover sheet glass 9, hydraulic control constant temperature sheet glass 10) wants high.Because the top alignment, preceding four bottom forms four depressions than both depressions of back 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 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.Corresponding six the porose rectangular preiections (Figure 15) of bottom flow turnover rectangular opening with top combination functional zone three hydraulic control constant temperature sheet glass 10 are arranged, and six circular holes at its corresponding back side and extraneous fluid pump into and pump pipeline be connected (Figure 16).Connection Block 2 main effect in bottom is to connect outside constant temperature fluid, support fixation entire structure, and being connected and fixed of convenient and 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.The both is inserted in the corresponding porose rectangular preiection on the side of fluid turnover sheet glass 9 of top combination functional zone (Fig. 9), and its position can be changed according to the actually operating demand.Fluid choke plug 4 makes the fluid in the corresponding porose rectangular preiection can not 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, promptly obtained passing through the finished product of designated period PCR round-robin reaction.
Described central fixed block 6 is a rectangular parallelepiped as shown in figure 13, and end face and bottom surface are squares.Material is an ABS resin.It is assembled in above-mentioned combination functional zone, top 1 and finishes when inserting described bottom Connection Block 2, can be stuck in bottom 2 four protruding blocks of Connection Block just and surround in the central zone that forms (Figure 15).The major function of central authorities' fixed block 6 has been central supporting roles 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, and the notch cuttype structure on its both sides is for corresponding with the notch cuttype structure of structure lateral column 11, and convenient location is steady with card during assembling.The major function of heat insulation rubber plate 7 is to make the removable micro-fluidic sheet glass 8 temperature homoiothermy that are in direct contact with it keep better, and it is too big that temperature is unlikely to be subjected to extraneous convection current and heat exchange to influence.Another function is to cushion as the structure fixed.
Described removable micro-fluidic sheet glass 8 is made by silica glass.Global shape is the rectangle thin slice, 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 bigger a little porthole (Fig. 7) of inner other passages, can with the porose projection corresponding (seeing Figure 11) on the side of the micro-fluidic sheet glass 8 of structure lateral column 11 clampings, make both fluid channels to engage and do not reveal.The fluid channel is a multiple tracks, and interbody spacer is opened mutually, 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 with Fig. 6.The use of can corresponding when reagent react, need under 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 mobile home court institute.
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 sheet glass 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 makes 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, promptly finish a circulation and enter next round-robin passage.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 make whole system reaction can access needed PCR cycle index, 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 promptly 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 sheet glass 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 inserts extraneous constant temperature water tank and follows constant temperature water tank.The thermostat(t)ed water that the outside pump housing pumps into will make hydraulic control constant temperature sheet glass 10 keep constant temperature.The 10 main effects of hydraulic control constant temperature sheet glass are exactly by thermal contact conductance, keep the directly temperature of the removable micro-fluidic sheet glass 8 of contact, make the PCR reaction obtain the temperature that needs.
Described structure lateral column 11, its structure such as Figure 11, shown in Figure 12.Wherein the structure on the both direction (being 90 ° of C mutually) 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 sheet glass 10 held.As shown in figure 11, porose projection on the medial surface has the mutual anastomosing fluid channels of two porose projectioies (Figure 12) in the horizontal plane in the porose projection.Corresponding to removable micro-fluidic sheet glass 8 or hydraulic control constant temperature sheet glass 10 lateral portholies.Can when assembling, the fluid channel well be sealed, make the fluid channel of the sheet glass that structure lateral column 11 two ends are followed connect.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.
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 same horizontal plane.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), each all is close to a less side side of central fixed block 6, 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 the one side by heat insulation rubber plate 7 of the slot of per two structure lateral columns 11 formation respectively.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, and the side that contains porose projection outwardly.3 blocks of hydraulic control constant temperature sheet glass 10 are inserted into the slot on 3 removable micro-fluidic sheet glass 8 next doors respectively then.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 sheet glass 10 all 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 sheet glass 10 or the structure lateral column 11 fluids turnover sheet glass 9 folded with it can be completely fixed non-loosening.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, screw is torqued into, functional zone 1 are made up on top promptly be completely fixed non-loosening with bottom Connection Block 2 from the screw column (seeing Figure 16) of bottom Connection Block 2 back.Top fluid control tube 5 has two screw holes, and its two screw holes with fluid turnover sheet glass 9 tops in portion 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.As Fig. 1, shown in Figure 180, combination functional zone 1, top promptly 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 sheet glass 10 is plugged pipeline, be connected to the outside pump housing and constant temperature water tank.Make the steady temperature of hydraulic control constant temperature sheet glass 10A, 10B, 10C be set at 94 ° of C(sex change respectively).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 the outside source of the gas or the pump housing.The external air source or the pump housing will make 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 formed of internal fluid channels flow circulation passage.According to 9-〉11A-〉8A-11B-〉8B-〉11C-8C-11D-9-... sequential loop flow, can pass in and out sheet glass 9 at fluid then and enter next circulation, wholely constitute a volution stream.Because removable micro-fluidic sheet glass 8A, 8B, 8C respectively with hydraulic control constant temperature sheet glass 10A, 10B, 10C directly fits, also can the flow through warm area of corresponding hydraulic control constant temperature sheet glass 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) order of warm area flows, and finishes 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 because preceding 4 rectangles are porose by fluid choke plug 4, thus branch road to go out head piece inoperative.The 25th round-robin branch road goes out head piece (corresponding the 5th porose projection of rectangle), and outside what connect 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 25 PCR circulation backs, finishes whole 25 round-robin PCR reaction process.
To sum up, the present invention proposes a kind of functional module that is used for the field-replaceable unit of pcr amplification, with removable micro-fluidic sheet glass is the principal reaction zone, the constant temperature fluid that pumps into the external world arrives hydraulic control constant temperature sheet glass as each warm area thermal source, carry out the reagent exchange by the fluid turnover sheet glass and the external world, with the structure lateral column of the fluid channel of four transfers for fixing, the center is useful on the 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.Upward convenient aligning of modular structure design and cooperation are fixing, fix whole each parts with 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 a PCR reaction efficiency height, 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 (10)
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 sheet glass (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 structure lateral column (11) is being fixed in central fixed block (6) corner, 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 sheet glass (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).
2. the functional module that is used for the field-replaceable unit of pcr amplification according to claim 1, it is characterized in that: 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 sheet glass (10) and structure lateral column (11), the top of these parts is on same horizontal plane, the bayonet socket of structure lateral column (11) is fitted fully with the corner of central fixed block (6) respectively, each piece all is close to a less side side of central fixed block (6) 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 the one side by heat insulation rubber plate (7) of the slot of per two structure lateral columns (11) formation respectively; 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 sheet glass (10) is inserted into the slot on removable micro-fluidic sheet glass (8) next door respectively.
3. the functional module that is used for the field-replaceable unit of pcr amplification according to claim 1 and 2, it is characterized in that: 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, the assembling back forms a spiral type stream, and fluid passes in and out sheet glass (9) turnover stream by stream.
4. the functional module that is used for the field-replaceable unit of pcr amplification according to claim 1 and 2, it is characterized in that: there is the fluid channel described hydraulic control constant temperature sheet glass (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 sheet glass (10) by the external world.
5. the functional module that is used for the field-replaceable unit of pcr amplification according to claim 1 and 2, it is characterized in that: described hydraulic control constant temperature sheet glass (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 a level, connects on two glass-chips the fluid with one deck; Flow out from following one deck outlet after then just in time making fluid flow into from this layer inlet fluid channel in the fluid turnover sheet glass (9).
6. the functional module that is used for the field-replaceable unit of pcr amplification according to claim 1 and 2, 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 projectioies 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.
7. the functional module that is used for the field-replaceable unit of pcr amplification according to claim 1 and 2, 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 that is raised with is individual, 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.
8. the functional module that is used for the field-replaceable unit of pcr amplification according to claim 1 and 2, 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 constitutes, 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.
9. the functional module that is used for the field-replaceable unit of pcr amplification according to claim 1 and 2, it is characterized in that: described top fluid control tube (5) top is the flexible pipe that can be connected with the 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 inlet of volution stream.
10. the functional module that is used for the field-replaceable unit of pcr amplification according to claim 1 and 2, 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 repeatedly round-robin PCR reaction, finally derive from outlet pipe (3).
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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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| CN102296027B CN102296027B (en) | 2013-04-17 |
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| 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 |
| 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 |
| CN107703320A (en) * | 2017-10-27 | 2018-02-16 | 大连量子流体控制技术有限公司 | Fully integrated Multi-channel multifunctional microfluidic analysis experimental system |
| 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 |
| CN109030437A (en) * | 2018-07-04 | 2018-12-18 | 中山出入境检验检疫局检验检疫技术中心 | A kind of detection device and detection method of 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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| 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 |
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| CN107703320A (en) * | 2017-10-27 | 2018-02-16 | 大连量子流体控制技术有限公司 | Fully integrated Multi-channel multifunctional microfluidic analysis experimental system |
| 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 |
| CN109030437A (en) * | 2018-07-04 | 2018-12-18 | 中山出入境检验检疫局检验检疫技术中心 | A kind of detection device and detection method of 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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