CN104741159B

CN104741159B - Programmable microfluidic digital array

Info

Publication number: CN104741159B
Application number: CN201510158081.2A
Authority: CN
Inventors: 布赖恩·福勒
Original assignee: Fluidigm Corp
Current assignee: Standard Biotools Corp
Priority date: 2008-12-08
Filing date: 2009-12-07
Publication date: 2017-01-18
Anticipated expiration: 2029-12-07
Also published as: SG172015A1; EP2376227A4; CN102281950A; SG10201404683TA; CN104741159A; SG10201404682WA; SG10201404685PA; WO2010077618A1; CN105964316A; EP2376227A1; CN102281950B

Abstract

The invention provides a microfluidic device, and more particularly, the present invention relates to a programmable microfluidic digital array and improved methods and systems associated with the microfluidic device. The microfluidic device includes a pressure source and a control line in fluid communication with the pressure source. The microfluidic device also includes a plurality of valves operated via the control line and an independent valve positioned adjacent the control line and between the pressure source and the plurality of valves.

Description

Programmable microfluidic digital array

The application is the divisional application of the Chinese patent application of Application No. 200980154761.x, and original application is 2009 The pct international application pct/us2009/067037 that December 07 submitted to enters the Shen of National Phase in China on July 15th, 2011 Please.

Technical field

The present invention relates to Programmable microfluidic digital array.

Background technology

Microfluidic device can be used for analyzing on the yardstick that still cannot imagine very recently, prepares, measures and other behaviour Make function.The advantage of microfluidic device includes saving the reagent of preciousness and the high density of sample, sample analysis or synthesis and high pass It is right that fluid degree of accuracy in amount, visually almost invisible level and accuracy and replacement operate on macrofluid yardstick The space that equipment is brought is answered to reduce.Reduce to microfluidic device size and density increase related be higher complexity and more Big engineering Manufacturing cost and finer device construction.

Although having these progress in microfluid design and application aspect, reduce complexity and the letter of micro-fluid chip Change operating or useful of they.Furthermore, it is necessary to enhanced control the correlation occurring in flow of fluid and microfluidic device The ability of course of reaction.Therefore, this area needs the improved method and system related to microfluidic device.

Content of the invention

The present invention relates to microfluidic device.More particularly it relates to Programmable microfluidic digital array and its operation Method.It is only used as illustrating, methods described and equipment have been applied in the system provide in micro-fluid chip asynchronous logic function. Additionally, check valve is integrated into digital array to provide lock-in control line by some embodiments of the present invention.However, it is recognized that The present invention has the much broader scope of application.

According to one embodiment of the invention, there is provided microfluidic device.Described microfluidic device comprise pressure source and Control line with pressure fluid communication.Described microfluidic device also comprises multiple valves operating by described control line, also wraps Containing separate valves, described separate valves are located at described control line nearby and between described pressure source and the plurality of valve.

According to another embodiment of the invention, there is provided operation has the side of the microfluidic device of valve and control line Method, described control line has coupled valve group.Methods described includes closing described valve and applying pressure to described control line. The valve of described cut out leads to the valve group being connected with described control line inoperable.

According to yet another embodiment of the present invention, there is provided microfluidic device.Described microfluidic device comprise the first valve, Second valve and the control line being in fluid communication with described first valve and the second valve.Described microfluidic device also comprises and described control line Be in fluid communication acute build up of pressure device (pressure accumulator) and be located at described acute build up of pressure device and described second valve Between described control line near check valve.

An alternate embodiment according to the present invention, there is provided microfluidic device.Described microfluidic device comprises many The individual reative cell with array layout arrangement.Each of the plurality of reative cell has and one of multiple reative cells fluid communication The first valve and with the plurality of reative cell in described in one of be in fluid communication the second valve.Described microfluidic device also comprises can Operation with start described first valve and described second valve the first control line and with the plurality of reative cell be in fluid communication defeated Enter line group.Described microfluidic device is also comprised the multiple sample inlets being in fluid communication with described input line group and is arranged in described Check valve in first control line.

Another alternate embodiment according to the present invention, there is provided microfluidic device.Described microfluidic device comprises Multiple reative cells and multiple first input port.Each of the plurality of first input port passes through one of more than first input line and institute State one or more fluid communication of multiple reative cells.Described microfluidic device also comprises multiple second input ports.The plurality of Each of second input port passes through one or more fluid communication that one of more than second input line is with the plurality of reative cell.Institute State the first pressure accumulator that microfluidic device also comprises to be in fluid communication with the first control line.Described first control line is set to off Close described more than first input line.Additionally, the second pressure that described microfluidic device comprises to be in fluid communication with the second control line stores Long-pending device.Described second control line is set off described more than second input line.In addition, described microfluidic device comprises to be arranged in The first check valve in described first control line between described first pressure accumulator and described more than first input line, and cloth Put the second check valve in described second control line between described second pressure accumulator and described more than second input line.

According to another alternate embodiment, there is provided operation has the microfluidic device of multiple valves and check valve Method.Methods described includes applying first fluid pressure to the control line of described microfluidic device, and responds to described first The applying of pressure and close the plurality of valve.Methods described also includes responding to the applying of described first pressure and closes described list To valve, and apply second fluid pressure to the second control line of described microfluidic device.

According to one embodiment of the invention, there is provided operation has the side of the microfluidic device of multiple input ports Method.Methods described includes providing input fluid to one of the plurality of input port, and start valve group with close with the plurality of The Part I input line that the subgroup of input port connects.Described subgroup does not comprise the one of described of the plurality of input port.Described Method also includes making described input fluid to flow through the input line being connected with one of described in the plurality of input port, makes described input Fluid flows through described input line and reaches the Part II of described input line and close the second valve group to separate multiple reative cells.

Another specific embodiments according to the present invention, there is provided the method for operation programmable microfluidic device, described Device has the reaction site array being in fluid communication with the first input line group and the second input line group.Methods described includes starting can Operation is in order to blocking the first valve group of described first input line group, and starts the operable Asia in order to block the second input line group Second valve group of the Part I of group.Methods described also includes loading sample by the Part II of described second input line group Enter described reaction site and start operable the 3rd valve group in order to separate described reaction site.

Another specific embodiments according to the present invention, there is provided microfluidic device.Described microfluidic device comprises pre- The input port of fixed number amount and multiple input fluid line, described input port each operable to receive multiple input one of fluids, Each of the plurality of input fluid line and one of the input port of described predetermined quantity fluid communication.Described microfluidic device also wraps Containing valve group, described valve group each operable to close one of the plurality of input fluid line.The quantity of described valve group is less than institute State predetermined quantity.Described microfluidic device also comprises each manifold being in fluid communication and the second valve with described input fluid line Group, each operable part with the described manifold of closing of described second valve group.

According to one embodiment of the invention, there is provided the method for operation programmable microfluidic device, described device has There is reaction site array, it is with the first input line group, the second input line group and the manifold stream being connected with described second input line group Body connects.Methods described includes starting operable the first valve group in order to close described first input line group, starts operable use To close the second valve group of the Part I of the subgroup of the second input line group, and start and be operable to make described manifold stop The 3rd valve group with (deactivate).Methods described also includes revocation and starts (deactuate) described second valve group, will be multiple Sample is loaded into described reaction site by the Part II of described second input line group, and start be operable to separate 4th valve group of described reaction site.

According to another embodiment of the invention, there is provided the method for operation programmable microfluidic device, described device There is the reaction site array being in fluid communication with the first input line group and the second input line group.Methods described includes starting can be grasped Effect is blocking the first valve group of described first input line group, and starts and be operable to separate the of described reaction site Two valve groups.Methods described also includes revocation and starts described second valve group, and multiple samples are loaded into institute by the second input line group State reaction site and start described second valve group.

According to yet another embodiment of the present invention, there is provided microfluidic device.Described microfluidic device comprises multiple anti- Answer site and the first input line group, described first input line group provides the first input port of predetermined quantity and the plurality of reaction position Fluid communication between point.Described first group of quantity is described predetermined quantity.Described microfluidic device also comprises the second input Line group, it provides the fluid communication between the second input port of predetermined quantity and the plurality of reative cell.Described second input line Each of group comprises trunk portion and component, and described second group of quantity is less than described predetermined quantity.Described miniflow Body device also comprises programmable input equipment, its operable in order to using described first input line group or described second input line Group loads described reative cell.

According to one embodiment of the invention, there is provided setting has the side of the microfluidic device of multiple control lines Method.Methods described includes starting the first control line and valve is placed in first state.Methods described is started after also including can quilt Operation in order to be placed in the second control line of the second state by valve group.The valve being in described first state stops described valve group from being placed in Second state.

Another specific embodiments according to the present invention, there is provided setting has the microfluidic device of multiple control lines Method.Methods described is included by starting the first control line with then starting the second control line sets up described microfluidic device First state, and by starting described second control line and subsequently starting described first control line and set up described microfluidic device The second state.

A kind of another specific embodiments according to the present invention, there is provided microfluidic device.Described microfluidic device bag Containing the first valve and the second valve.Described microfluidic device also comprises the control being in fluid communication with described first valve and described second valve Line, and the pressure source being in fluid communication with described control line.Described microfluidic device also comprises to be arranged in institute in described control line State the check valve between pressure source and described second valve.

Another alternate embodiment according to the present invention, there is provided microfluid system.Described microfluid system comprises Carrier.Described carrier comprises multiple first input ports and multiple first input line.Each and institute of the plurality of first input line State one of multiple first input ports to be in fluid communication.Described carrier also comprises multiple second input ports and multiple second input line.Institute Each stating multiple second input lines is in fluid communication one of with the plurality of second input port.Described carrier also comprises first pressure Source and second pressure source.Described microfluid system also comprises to be contained in the microfluidic device on described carrier.Described microfluid dress Put and comprise multiple 3rd input lines and multiple 4th input line.Each of the plurality of 3rd input line is defeated with the plurality of first Enter one of line to be in fluid communication, each of described 4th input line is in fluid communication one of with the plurality of second input line.Described micro- Fluid means also comprises the first control line with described first pressure fluid communication, is operable to block described first control At least one of check valve of line processed, and the second control line with described second pressure fluid communication.

The present invention has the many advantages surmounting routine techniquess.For example, the technology of the present invention allows to customize after fabrication miniflow Body device, allows users to provide the setting of multiple programmable panels.These and other embodiments of the present invention and its permitted Many advantages and features contacts are hereafter described in more detail with accompanying drawing.

Brief description

Fig. 1 is the rough schematic view of microfluidic device according to an embodiment of the invention.

Fig. 2 a is the rough schematic view of check valve according to an embodiment of the invention.

Fig. 2 b is the simplified top view showing check valve shown in Fig. 2 a.

Fig. 3 is a kind of simplified flowchart of operation method of microfluidic device according to an embodiment of the invention.

Fig. 4 is a kind of simplified flowchart of operation method of microfluidic device according to another embodiment of the invention.

Fig. 5 is the simplified flowchart of a kind of method according to the microfluidic device of further embodiment of the present invention for operation.

Fig. 6 a is the sketch of reative cell in array according to an embodiment of the invention.

Fig. 6 b is that the simplification of control line, fluid input line and reative cell in array according to an embodiment of the invention is saturating View.

Fig. 7 shows a kind of method for simplifying to programmable microfluidic device programming according to an embodiment of the invention.

Fig. 8 shows a kind of simplification side to programmable microfluidic device programming according to another embodiment of the invention Method.

Fig. 9 shows a kind of simplification side to the programmable microfluidic device programming according to further embodiment of the present invention Method.

Figure 10 is the rough schematic view of programmable microfluidic device according to an embodiment of the invention.

Figure 11 is the rough schematic view of microfluid system according to an embodiment of the invention.

Specific embodiment

Fig. 1 is the rough schematic view of the microfluidic device according to one embodiment of the invention.In a specific enforcement In scheme, the microfluidic device shown in Fig. 1 comprises Programmable high-density digital array.Described microfluidic device 100 comprises first Pressure source 102 and second pressure source 104.In FIG, first pressure source 102 is referred to as accommodating accumulator (containment Accumulator), as described in more detail below, described pressure source 102 and control line are in fluid communication, and described control line is operable In order to close the valve being associated with reative cell (not shown) in array 106.Because starting of being carried out using pressure source 102 makes these Reative cell valve cuts out and makes sample and/or reagent to be contained in described reative cell, therefore pressure source 102 in some embodiments Referred to as accommodate accumulator.

In operation, liquid is placed in acute build up of pressure device (pressure accumulator) or pressure source, described Acute build up of pressure device or pressure source are subsequently connected with outside malleation supply (as the container containing compressed air).Under stress, described Liquid in described acute build up of pressure device is pushed control line, thus starting valve by compressed air or other fluids.Therefore, described pressure Source does not generally comprise pressure fluid during fabrication, and during being to provide for operating microfluidic device, pressure fluid can be accumulated wherein With storage so that upon activation control line is applied with stressed container.As being described more fully in text of the statement, described pressure Accumulator operable to maintain the pressure in control line upon activation.In CO-PENDING and commonly assigned U.S. Provisional Patent Application No.61/044, provides the additional description of the digital array being suitable for carrying out embodiment of the present invention in 417, in its entire disclosure Hold and all purposes is passed through with overall being incorporated herein by reference.

Fig. 6 a is the sketch of reative cell in array according to an embodiment of the invention.As an example, the present invention Some embodiments using the cell with reative cell, its lateral dimension be 60 μm of 100 μ m, highly for 125 μm.At this In exemplary, the volume of described room is about 0.75nl.So little building volume can using less sample volume simultaneously Decrease running cost.Through hole (via) the coupled reaction room that 50 μm of diameter and mensure/sample input line.With 110 μm of the first side The second lateral room spacing placing response room to room spacing and 200 μm.In the layer different from the layer containing mensure/sample input line The middle valve providing 50 μm of 50 μ m, described valve is operable to flow through described input line in order to stop.Outside the particular geometric of this cell Shape is not intended to limit embodiment of the present invention, and simply provides the example of specific embodiments.In another embodiment, Using other device geometric shapes being adapt to application.

According to some embodiments of the present invention, using width range be about 5 μm to about 400 μm, depth bounds be about 5 μm The fluid flowing through described microfluidic device is provided to about 75 μm of input lines.Width range is about 5 μm to about 400 μm, depth model Enclose be about 5 μm to about 75 μm control line be used for by valve close (valve off) through input line flow of fluid.At some In embodiment using width range be about 10 μm to about 500 μm, length be about 10 μm to about 500 μm, altitude range be about 5 μm To about 500 μm of reative cell.The geometric shape of these devices provides by way of example, and is not intended to limit described herein Embodiment.

As shown in Figure 6 a, there is provided multiple input lines 620, fluid is enable to flow through described input line in the horizontal direction.Display Three input lines, but some embodiments of the present invention are using the input line more than three, for example, 11 input lines.Using Then single sample can be distributed in multiple input lines a plurality of input line, and this provides multiple copies of given sample.As to Fig. 6 b Described in the more complete description carrying out, described input line or input channel are at least partially embodied in the first of microfluidic device In layer.Referring to Fig. 1, described input line is in fluid communication with the mensure input line 132 in array 106 left side, and with array 106 right side Sample input line 142 is in fluid communication.Therefore, all it is provided to input from the fluid measuring input 130 or sample input 140 Line simultaneously and then is supplied to reative cell.

There is provided multiple control lines 610 in vertical direction, enabling control the flow of fluid through input line.Show Article two, input line, but some embodiments of the present invention are using the control line more than two, for example, 70 control lines.Another In individual embodiment, there are 71 control lines.Described a plurality of control line forms detached reative cell along the length of input line, provides Comprise multiple reative cells of same sample.As described in the description more complete to Fig. 6 b, described control line or control passage are extremely Small part is included in the second layer of microfluidic device.Referring to Fig. 1,114 part streams of described control line and lock-in control line 110 Body connects, and described lock-in control line is in fluid communication with accommodating accumulator 102.Intersection point shape in described control line and described input line Become valve 615, it responds to the Fluid pressure in control line and actuated, and operable with stop fluid flow through described input line. Generally, described multilamellar microfluidic device comprises multiple elastomer layers and valve 615 comprises deflectable diaphragm.Show in Fig. 6 a and 6b In embodiment, the deflectable diaphragm of described valve deflect to be located at the fluid passage on the intersection point of control passage in.Therefore, institute The embodiment of display adopts " above pushing away " valve, and wherein deflectable diaphragm upward deflects to fluid passage to close in valve position Fluid passage.For the valve of display in Fig. 6 a and 6b, Fluid pressure present in release control passage leads to deflectable diaphragm to be returned Return undeflected position thus the valve of either on or off.

The fluid flowing through input line 620 through through hole 625 and flows upwardly into reative cell along the direction vertical with Fig. 6 a plane 630, as reative cell 630 as to described in being described more fully of Fig. 6 b is at least partially embodied in the 3rd of microfluidic device In layer.Therefore, described through hole is at least partially embodied at least described second or third layer of microfluidic device.Generally, adopt Laser ablation process remove second or third layer a part of forming described through hole.Because described microfluidic device is that air can Through, using closing filling technique (blind fill technique) filling reative cell and can carry out multiple chemical, biological Or other experiments.Skilled in the art realises that, after there is fluid in the reaction chamber, starting of control line will lead to valve to be closed simultaneously Fluid is retained in the reaction chamber predetermined time period.

Fig. 6 b is the simplification perspective of the control line, fluid input line and reative cell in the array of one embodiment of the invention Figure.Array (for example, the array 106 shown in Fig. 1) is a part for multilamellar microfluidic device.Every layer generally comprise have one or The elastomer structure of more grooves, passage, room etc..As shown here, ground floor 601 comprises to be arranged to array of parallel channels Multiple control passages 610 and other control passage 611.Described control passage 610 and other control passage 611 and one or More pressure sources or acute build up of pressure device are in fluid communication.Therefore in one embodiment, control passage 610 and acute build up of pressure device 102 fluid communication, control passage 611 is being in fluid communication with acute build up of pressure device 104.Although showing that single control is led in figure 6b Road 611, but those of ordinary skill in the art are it should be understood that shown single passage represents being associated with 601 layers or more Multiple control passages.

Additionally, described control passage is not restricted to the position related to layer 601.Described control passage is adaptable to specifically should With and be placed in other layers.For example, in the enforcement of a multiple control (control-on-control), in order that the first control Line (such as control passage 611) to the second control line (such as control passage 610) produce control, by described second control line according to The length of control passage is placed in both ground floor 601 and the second layer 602.Using the through hole similar to through hole 625 by second Control line is transformed into the second layer 602 from ground floor 601.By passing through the second control line above the first control line, control at two The intersection point of line processed forms valve.Once starting described first control line, then between described first control line and described second control line The flexible membrane of valve position upwardly to block the second control line in being located at 602 layers in valve position.Therefore, as hereinafter more Complete description, " above pushing away " valve can be formed between different control lines or between control line and fluid input line.Referring to Fig. 1, with The control line that valve 128 is connected is transitioned into upper strata by through hole from the lower floor of structure, then makes control line 122 through control line 122 So as to start separator valve 124.Then, the control line being associated with valve 128 wear through another through hole return in ground floor from And pass through and start the valve of fluid input line 132.Once control line 122 actuated, separator valve 124 obstruction passes through and valve 128 phase The flowing of control line even, stops valve 128 from generally responding to the closing that control line 110 starts.

The second layer 602 comprises also with multiple fluid inlet channel 620 of array of parallel channels arrangement.Shown in Fig. 6 a and 6b In the embodiment shown, control passage 610 and input channel 620 arrange perpendicular to each other.By existing in control passage 610 Liquid (usually substantially incompressible fluid) apply pressure realizing starting of valve present in microfluidic device.Logical Often, liquid is placed in accumulator (accumulator) or pressure source and provides the fluid of pressurization (for example empty to accumulator Gas, nitrogen etc.).In accumulator, the increase of pressure makes liquid enter control line under stress.Respond to pressure applied, formed The deflectable diaphragm at 601 layers of top upward deflects entrance fluid inlet channel 620.Therefore, in control passage and fluid inlet channel Intersection point produces " above pushing away " valve.Some other embodiments can by reorientate various control and fluid layer using " under push away ", " under Draw " or " pull-up " valve.

In some embodiments of the present invention of multiple control are provided, it is possible to provide one or more other key-courses (such as lower floor 601), or by control line is arranged into 602 layers.One or more other key-courses can comprise Other control line (not shown), it once starts by control line present in valve closed layer 601.Therefore, by one Fluid (such as liquid) is controlled to apply Fluid pressure present in individual or more other control lines, flexible membrane deflects to control Line 610, stops fluid from flowing through control line.Therefore, some embodiments of the present invention provide key-course, also referred to as multiple control System.Those of ordinary skill in the art are it will be appreciated that many change, improve and replace.

The described second layer also comprises multiple through holes 625, produces and provides from described input channel to the fluid stream of third layer 603 Dynamic fluid passage.Third layer 603 comprises multiple reative cells 630, and it is passed through through hole and is in fluid communication with described fluid inlet channel. In the embodiment illustrated, reative cell 630 is formed as being connected with the bottom of layer 603, and in other words, described reative cell from bottom is Open.Therefore, through hole is completely contained in 602 layers.In other embodiments, described through hole can be comprised in 602 With 603 layers in both to be adapted to specific application.

In some embodiments, microfluidic device can comprise according to rotation or cast fabrication scheme preparation one or more Multiple layers.For example, rotation approach can relate to place polymeric material on figuratum disk or mould, and rotates described disk with disk Upper generation polymeric layer.Illustrative polymers include polymethacrylates, polystyrene, polypropylene, polyester, fluoro polymerization Thing, politef, Merlon, polysilicon and polydimethylsiloxane (polydimethylsiloxane, pdms).Cast Scheme can relate to as being cast in pdms material in figuratum template or mould, and this can produce and intactly can peel from mould Or the pmds layer left behind.Generally, by pouring into a mould the layer of manufacturing technology preparation than the thickness with rotation manufacturing technology preparation.Elastomer block One or more cast or the rotating layer of any required combination can be comprised.

In some embodiments, ground floor 603 can be manufactured according to rotation approach.For example, pdms can be placed on and have On mould corresponding to the bossing of multiple required control passages 610.Rotatable described mould is to produce on whole mould Raw pdms thin layer.After hardening, ground floor 601 can be peeled off from mould and be attached to suitable rigid matrix (as glass, silicon) Or on plastics (polystyrene).Or, ground floor 601 can remain and be attached on mould.Ground floor 601 can comprise opening, recessed Groove or other spaces being at least a partially formed or limiting control passage 610.

In order to generate the second layer 602, rotation approach can be adopted, pdms is placed on the second mould, described second mould There is the bossing corresponding to multiple required fluid inlet channel.Second mould also can comprise, for example, formed corresponding to second Projection or the wavelike segmentss of labelling are arranged in layer 602.These arrangement labellings can make in the laser ablation for forming through hole 625 With so that laser ablation points to described arrangement labelling during ablation processes.Rotatable second mould is with whole second mould Pdms thin layer is provided.The second layer 602 can comprise opening, groove or be at least a partially formed or limit other skies of input channel 620 Gap.In some cases, the second layer 602 can be exposed to the above-mentioned laser ablation of one or many.Point to the ablation of the second layer 602 Laser beam can form through hole 625.After the second layer 602 fully solidifies, ground floor 601 can be peeled off from the second mould, and by its Align with ground floor and contact.The second layer can be pasted so that two-layer sticks on rigid matrix with ground floor.

In order to third layer 603 is produced using casting program, pdms can be cast in and have corresponding to multiple required reactions On 3rd mould of the bossing of room 630.After hardening, can peel off from the 3rd mould third layer 603 and with the second layer 602 Align and contact.Third layer can be pasted on the second layer so that all three layers stick on rigid matrix.Microfluid can be manufactured The material of device includes but is not limited to elastomer, silicon, glass, metal, polymer, pottery, inorganic material and/or these materials Combination.

Referring back to Fig. 1, pressure source 102 is in fluid communication with closing control line 110.Described lock-in control line comprise several hereafter The section being described more fully and check valve 112 (also referred to as check-valves (check valve)).In CO-PENDING and commonly assigned International patent application no.pct/us07/080489 (, its entire disclosure open with international publication number wo 2008/043046 a2 Content passes through overall being incorporated herein by reference) in provide other description with regard to check valve.First section 114 is from pressure source 102 Reception start pressure and operable in order to close present in closed array 106 accommodate valve (containment valve).As this area Technical staff is understood ground, and array 106 is adapted for multiple micro-fluid experiment.Thus can be by responding to lock-in control line 110 Start and close and accommodate valve so that sample, reagent etc. present in reative cell in array retain in the reaction chamber.Locking control The section 114 no check-valves of line 110 processed, this make receiving valve remove pressure source 102 start pressure after reopen.

Second section 116 of lock-in control line 110 is in the downstream of check valve 112.Due to check valve 112 operable in order to hinder Fluid stopping body flows to section 114 from section 116, and starting of pressure source 102 will lead to control fluid (usually liquid) by unidirectional Valve 112 shutoff valve 118a-118e.The fine rule being associated with lock-in control line 110 shown in Fig. 1 represents the " high of control line Frame (flyover) " section, the design of wherein control line prevents mensure input line 132 (being described more fully hereinafter) from closing Lock control line 110 is closed after starting.The thick line being associated with valve 118a-118e is represented and is operable to close or disables manifold The valve of 135 (providing in multiple flow of fluids measuring between input line 132).Close to stop and measure stream in the pass of valve 118a-118e Measure input line flow direction other mensure input line through manifold 135 from one, this will be referred to as the deactivation of manifold.

After starting lock-in control line 110 by pressure source 102, valve 118a-118e can close and remove locking control Line processed start pressure after, check valve 112 can keep the closed mode of valve 118a-118e.Contrary with valve 118a-118e, by area The receiving valve that section 114 is started can be opened after starting pressure described in removing.Therefore, in lock-in control line check-valves locus Provide the one or more sections being characterized with locking or non-blocked behavior.As those skilled in the art clearly, in control The extra section with locking behavior can be provided using extra check-valves in line processed.

Lock-in control line 110 also comprises the 3rd section 128, its have be operable to close from measure input 130 to The valve of the mensure input line 132 of array 106.Measure input 130 and may be additionally referred to as input port.Because the 3rd section 128 is also in list To the downstream of valve 112, lock-in control line 110 is started by pressure source 102 and can close the valve in the 3rd section 128 and stop in survey Determine the flowing between input line 132 and array 106 and the reative cell arranged thereon.Removing starting of lock-in control line 110 After pressure, the valve of section 128 remains turned-off.

There is provided the second pressure source 104 being in fluid communication with the second control line 120, it is referred to as interface accumulator (interface accumulator).Described second control line also comprises several sections 121,123 and 125.Section 121 comprises Elevated region 122 enables to start separate valves 124 in the case of not closing mensure input line 132.In this specification In provide another to valve 124 (referred to herein as independence or separator valve (independent or decoupling valve)) Outer description, and hereinafter it is described more specifically.Although described specific independence or separator valve 124 make in FIG Started with second pressure source 104, but this is optional in the present invention.In other embodiments, independence or separation Valve 124 can be mechanical, electrostatic, fluid, electromechanical, thermodynamic, piezoelectricity etc..Therefore, although with aobvious in Fig. 1 Separator valve 124 is started in the second pressure source 104 showing, but this is optional in some embodiments.Although additionally, using Single second pressure source 104 controls the multiple valve groups comprising separator valve, and other embodiments can adopt multiple pressure sources The combination that fluid starts and electrostatic start.Those of ordinary skill in the art understand multiple changes, improvement and replacement.

Section 123 comprises the valve 134 being operable to close 5 in the mensure input line 132 shown in 6.Due to 5 valves 134 are only provided, in the section 123 carrying out by pressure source 104, valve 134 starts still in embodiment shown in Fig. 1 The mensure input line that low order end can be made keeps it turned on.Section 125 comprises check valve 127 and is operable to close sample input The valve 126 of line 142, described sample input line 142 inputs (i.e. input port) 140 with sample and array 106 is in fluid communication.It should be noted that Although showing mensure and sample input in FIG, the invention is not restricted to only measure and sample input, other inputs include In the scope of the invention.

Unidirectional or check-valves 127 are placed on the section 125 of lock-in control line 120 it is allowed to 126 valves cut out and in section institute Remain turned-off after applying pressure, stop fluid from passing through the flowing of sample input line dealing array 106.Due to section 121 and 123 not Comprise check-valves, the revocation of these sections is started and will be led to reopening of valve 134 and 124.

Some embodiments of the present invention provide asynchronous logic function in microfluidic device 100.For example, due to using Individually and start the independence shown in Fig. 1 or separator valve 124, the present invention independent of the control line 120 of control line 110 shown in other Some embodiments provide multiple control.In FIG, before starting separator valve 124, pressure is applied to lock-in control line 110 Power locking can accommodate valve 128.On the other hand, before pressure is applied to lock-in control line 110 by receiving accumulator 102, pass through Interface accumulator 104 starts lock-in control line 120 thus closing separator valve 124 will stop the closing accommodating valve 128.In other words, If separator valve 124 cuts out when valve 128 is started apply with pressure, the closed mode of separator valve stops pressure applied from arriving Reach valve 128.Therefore, the order of starting of control line 110 and 120 leads to different valves to be operated, and this provides asynchronous logic.

The multiple control that embodiment of the present invention provides makes array 106 can be " programmed ".For example, because array 106 is pressed Multiple valves closings, locking or the sequential programming reopening, can use array 106 with multiple layouts.As herein described one In a little embodiments, provide three kinds of different layouts using 2 shown pressure sources.Using check valve and separator valve (its Can be considered one of stacking control valve group) these multiple layouts are provided.Separator valve can limit the control to valve 128, described control System can be by responding to the realization of starting of lock-in control line 110, and described separator valve is an example stacking control valve, this Bright scope includes other layouts.

Provide the part locking of control line in the predetermined segment of control line using check-valves.For example, can be by initial control Line 110 processed start locking-valve 118 and 128.However, initial control line 120 start the closing that can lead to separator valve.Subsequently Starting of control line 110 can lead to the locking of valve 118 still valve 128 no to be affected.Subsequently same keep control line 110 to start When revocation start after control line 120 (for example passing through to discharge the pressure that interface accumulator applies) can lead to separator valve to reopen The locking of valve 128.

In microfluidic device 100 shown in Fig. 1,6 measure input line 132 and are each separated into 4 input lines, there is provided wear Cross the 24 mensure input lines altogether accommodating valve 128.In addition it is shown that 24 sample input lines 142.These input lines concrete Numeral is example, and other embodiments provide other specific input line numbers.For example, in a specific embodiment, Schematic diagram shown in Fig. 1 illustrate only half microfluidic device (right side of such as device), has phase in the left side of described device The input group joined.Those of ordinary skill in the art are it will be appreciated that many change, improve and replace.

Fig. 2 is the simplification sectional view of the check valve of one embodiment of the invention.Fig. 2 b is the letter of check valve shown in Fig. 2 a Change top view.Referring to Fig. 2 a, start fluid (usually liquid) and flow into check-valves 112/127 through entrance 210, and through through hole 215 flow through room 220.By starting the flowing of fluid so that film 230 lifts or is maintained at the position of basic horizontal, make to start fluid From left to right pass through through entrance 210.Start fluid via through holes 240a-240d and through hole 242 flows through room 220 and from outlet 250 Flow out.Therefore, in the embodiment illustrated, for each check valve altogether 6 through holes, there is an input through hole and 5 Individual output through hole.As shown in the top view in Fig. 2 b, provide extra structural detail and assembly according to specific design.

When Fluid pressure is started in removal, film 230 is collapsed upon on layer 260, stops and is flowed back by entrance 210.Therefore, unidirectional Valve 124/127 provides the flowing from entrance 210 to outlet 250, but stops reverse flowing.Other with regard to check valve are described in There is provided in CO-PENDING and the commonly assigned above-mentioned application quoted.

Some embodiments of the present invention provide the microfluidic device being integrated with unidirectional or check-valves.In an embodiment party In case, described microfluidic device comprises the first valve and the second valve.Generally, described first valve is in the multiple valves formed the first valve group One, described second valve is one of the multiple valves forming the second valve group.With described first valve and described second valve fluid The control line of connection is provided together with pressure source as a part for microfluidic device, described pressure source and described control line fluid Connection.Referring to Fig. 1, by starting control line 110 to accommodating accumulator 102 and applying pressure, lead to valve 615 in array 106 with And valve 118a-118e starts so that manifold 135 disables.Described microfluidic device also comprises to be arranged in described pressure source and described The check valve in control line between second valve.For example, the valve 112 originally opened is located at and accommodates accumulator 102 and valve 118a- On control line 110 between 118e.After reducing or removing the pressure on control line 110, check-valves 112 provide valve 118a- The locking of 118e, and valve present in array 106 (non-blocked valve) can reopen and make sample flow into reative cell.Therefore, lead to Cross the valve using predetermined number in non-return valve blocking microfluidic device, there is provided irrealizable microfluidic device in being commonly designed The control of middle flow of fluid and the separation of sample.

In embodiment shown in Fig. 1 and 6a, valve 615 is arranged to reaction on microfluidic device for the spaced apart Room, and valve 118a-118e be arranged to by with measure input port 130 be connected (coupled) six lines first fluid defeated Enter line to separate with the second fluid input line phase measuring six lines that input port is connected.As more completely retouched in this manual State, close and locking-valve 118a-118e is so that the ability that manifold 135 disables enables microfluidic device to compile in many ways Journey, this enables the flow of fluid in input line 132 to be separated according to concrete application or to merge.

In addition to check-valves are integrated in the microfluidic device have array layout, some designs provide independence Valve, also referred to as separator valve.For example, independent of other valves in device controlled separator valve be operable to stop one or The starting of more valves, one or more valves control the fluid by being connected with reative cell present in microfluidic device The flow of fluid of input line.For example, separator valve 124 can be closed before starting control line 110, thus stoping valve 128 from responding to The closing that control line 110 starts.

Figure 10 is the rough schematic view of the programmable microfluidic device of one embodiment of the invention.Unit shown in Figure 10 Part can provide in the carrier or provide in the microfluidic device be attached at carrier as shown in figure 11.As shown in Figure 10, it is loaded on load The profile of the microfluidic device on body is represented by dotted line 1050.Referring to Figure 10, provide two in the opposite side of microfluidic device Emission source (vent source).The emission source providing usually as a part for carrier can not adopt in all embodiments. Show 48 sample input lines 1010 above Figure 10, the wherein left side in microfluidic device arranges 24 sample inputs Line simultaneously provides sample to array 106 left side, arrange 24 sample input lines on the right side of microfluidic device and to array 106 on the right side of Sample is provided.Article 48, sample wire generally provide in the carrier and by formed in microfluidic devices and with sample wire end pair Neat through hole 1030 and array 106 are in fluid communication.There is provided in microfluidic devices from through hole 1030 to the input of the sample of array 106 Line 142.Article 48, sample wire passes through the pressurization of UP source (common pressure source) (not shown), described pressure source The sample part of the carrier being placed on it with microfluidic device is connected.Therefore, the sample of 48 sample ports can be written into miniflow Body device simultaneously pushes array 106 through sample input line.Referring to Fig. 1, for the sake of clarity show the right side of array 106 Point, wherein the 24 of Figure 10 right part sample input line is shown as sample input line 142.Therefore, some enforcements of the present invention Scheme provides 48 sample input lines with up to 48 different samples.Other enforcements provide difference according to concrete application Number.

Show control line 1004 in Fig. 10, the interface accumulator 104 that it is shown with Fig. 1 is connected.Additionally, in Fig. 10 Show several check-valves (check valve, cv), this provides the one-way fluid flow through control line in microfluidic devices Dynamic.Referring to the bottom of array 106, provide other cv for connecting to the control line of this part of array.Outside in array 106 Provide aquation line.These aquation lines provide source for aquation fluid, and the effect of aquation fluid is to reduce or prevent in array The evaporation of fluid.

48 input lines showing in Figure 10 bottom comprise 12 and measure input line (6, array 106 left side, array 106 right side Side 6).Article 12, measure line and pass through the (not shown) pressurization of UP source, described pressure source can be placed it with microfluidic device On carrier mensure part connect.Therefore, 12 mensure mouths mensure can be written into microfluidic device and after measured input line push away Enter 2 arrays 106.Referring to Fig. 1, for the sake of clarity show right array 106, it has the figure being shown as measuring input line 132 6 sample input lines of 10 right part.Therefore, some embodiments of the present invention provide and have up to 12 different mensure 12 mensure input lines.Measure input line to pass through between two, the central authorities of microfluidic device array 106,6 lines are branched off into Right 106,6 lines of array are branched off into left array 106.The use of shown manifold is provided that and measures less than 12, for example, as little as single Measure.Other enforcements provide different numbers according to concrete application.

In other 36 input lines shown in Figure 10 bottom, 2 lines are used for aquation, and during here is embodied as, 34 is not Use.Measuring input line 1020 provides the input of the single input line entering array 106, and measures input line 1022 and be branched off into The a plurality of input line (for example, 4 lines) entering mensure 106.

Fig. 6 a shows the example of every 3 input lines of sample, but in the embodiment depicted in fig. 10, has 11 inputs Line.Additionally, in Fig. 6 a, 2 control lines are shown in Fig. 6 a, but in the embodiment depicted in fig. 10, there are 71 control lines 70 reative cell/input lines can be operable to form.Therefore, there are 770 reative cells for each sample in the present embodiment (11 input line × 70 reative cell).It is dispensed into 770 reative cells with 48 samples, there is provided highdensity integrated fluid Loop (integrated fluidic circuit, ifc).In one embodiment, microfluidic device energy one-time detection is many Reach 48 independent samples.Each of described up to 48 samples is dispensed into 770 reative cell independence groups, thus delivering as always The synchronous digital pcr reaction for 36,960 for the number.In another embodiment, some programmability of microfluidic device are removed, Then for identical device area and size of components, increased the reaction number of chambers/sample.For example, an embodiment is using up to 48 samples, it has 814 reative cell/samples.

Using the product being available from assignee of the present invention, entirely digital pcr mistake can be completed in less than 4 hours Journey.Additionally, microfluidic device as herein described is the ingredient of complete gene alaysis system.This system can comprise microfluid Device, the controller of microfluidic device, biomark^tmSystem or integral type thermal cycler, ep1 read instrument and related software.Herein Described microfluidic device is compatible with the distribution design of finished product reagent and cellular form.Those of ordinary skill in the art understand many Change, improve and replace.

Figure 11 is the rough schematic view of the microfluidic device of one embodiment of the invention.Described microfluid system comprises to carry Body 1100 and the microfluidic device 1108 being loaded on described carrier.Microfluidic device 1108 incorporates the element described in Fig. 1.Institute State multiple first input ports or the hole 1105 that carrier comprises to be arranged on platform (bank) 1106a.Described carrier also have with multiple Multiple first input lines 1115 (for example, 48 input lines) that first input port is in fluid communication.In the plurality of first input port Surrounding has outer peripheral edge, allows to UP source to described first input port pressurization.In an exemplary application, by 48 Sample is loaded into the first input port, from the top of carrier, described mouth is applied with pressure and leads to 48 samples to be pushed over multiple first inputs Line simultaneously eventually enters into the fluid line on microfluidic device.

Described carrier also comprises multiple second input ports 1105 being arranged on platform 1106b and multiple corresponding second defeated Enter line 1115.Similar to multiple first input ports, the plurality of second input port is by outer peripheral edge around making to be dispensed into the second input The fluid of mouth can be pushed over the second input line and enter the fluid line in microfluidic device.

It is in fluid communication to provide and the microfluidic device being placed on described carrier between in described carrier, microfluidic device There is the through hole 1114 being formed in microfluidic device bottom, its terminal part with described first input line and described second input line Divide alignment.The fluid flowing through input line is upwardly into the fluid line of microfluidic device offer through through hole.As shown in Figure 10, connect The fluid line of described sample input port is through entering in a subtle way with the through hole 1030 of Right vertical arrangement along on the left of in figure microfluidic device The offside of fluid means, and flow to the central authorities of microfluidic device.In Fig. 6 a, these sample input lines can level in corresponding diagram 6a The input line 620 flowing through.Using this 48 sample input lines, different samples can be filled in the reative cell that microfluidic device is often gone Product.

Additionally provide through hole 1040 in the bottom of microfluidic device and it is alignd with multiple second input lines 1020.As figure Shown in 10, the fluid (as measured) being loaded into the second input port flows through the second input line 1020, enters manifold through through hole 1040 1060.Described microfluidic device comprise can with described manifold crew-served valve, to lead to 1 fluid (sample or mensure) or 12 Individual different fluid is provided the reative cell in two arrays.If application needs single input fluid, manifold is unlocked simultaneously And input fluid flow in Figure 10 through array centre perpendicular through all fluid lines.Fluid line is through the array centre time-division Prop up away, the through hole 1030 finally flowing through array side with them finally flows through reative cell.Referring to Fig. 6 a, these measure input Line can correspond to the input line 620 passed through under reative cell.In this example, the flowing of left array side is from central authorities to left side, right The flowing of array is from central authorities to right side.

Or, can close manifold 135 to introduce the different fluid (for example measuring) of 12 reative cells to be introduced (is each battle array 6 different mensure of row).As shown in Figure 10, fluid perpendicular flow between array, then branch go out, wherein 12 fluids Each flow into array multiple rows of in, flatly pass through to the neighbouring through hole in edge 1030 from central authorities.

Carrier provide acute build up of pressure device 1106c and 1106d enable to start control present on microfluidic device Line processed and check-valves.With with fluid line similar mode, the control line 1002 and 1004 in carrier pass through microfluidic device on shape The through hole becoming is in fluid communication with the control line 110 and 120 on microfluidic device.Therefore, while figure 1 show that the interface of carrier and Accommodate the control line in accumulator and microfluidic device it should be understood that this schematic diagram is the sketch for the sake of clear and convenient.With The other details of carrier correlation that microfluidic device can be placed thereon are in U.S. Patent Application Publication no.2005/0214173 There is provided, the disclosure of which passes through overall being incorporated herein by reference for all purposes.

Carrier 1100 has the acute build up of pressure device hole 1101 and 1102 of integration, has the dry hole for accommodating valve in each of which 1103rd, 1104, described valve is preferably the check-valves being connected with lid.Carrier 1100 also comprises one or more hole platforms 1106a, b, c And d, each of which has the one or more holes 1105 (also referred to as input port) being located therein.The hole 1105 of each carrier 1100 has There is the passage guiding the microfluidic device 1108 being placed in carrier positions 1107 from hole 1105.Hole platform 1106c and 1106d are generally used for The pressure being used for starting the control line in the presence of microfluidic device 1108 is provided.There is provided the fluid line controlling fluid, by hole platform In hole be connected with valve present in microfluidic device or other control devices.Microfluidic device is preferably by two-layer or more The elastomer block that layer elastomeric material is formed, it has the groove of the micro manufacturing being formed wherein or passage.

The one or more passages being in fluid communication with one or more through holes 1114 are had, it is then in described microfluidic device There is provided the fluid communication between the passage in the passage and carrier in microfluidic device, then in its pilot hole row 1106a-d Hole 1105, thus provide the fluid communication between the passage in the hole 1105 and microfluidic device 1108 in carrier 1100.Accumulation Device hole top 1109 and 1110 is connected to accumulator bore 1101 and 1102 to form accumulator room 1115 and 1116.Accumulator bore top 1109 and 1110 comprise valve 1112 and 1111 respectively, and it preferably introduces a gas into accumulator room 1115 and 1116 under stress And retain the check-valves of gas.When being present in accumulator room 1115 and 1116, valve 1111 and 1112 is located at dry hole 1102 He 1104 inside is not so that liquid contacts valve 111 and 1112.Preferably by preferred check-valves by laminated flake (shave), mechanically opening valve 1111 and 1112 such as pin, it overcomes the power that is automatically switched off of check-valves to press to discharge from accumulator room Power, thus reduce contained Fluid pressure in accumulator room.

Carrier 1100 and its associated component can be by polymers manufacturings, and such as polypropylene, polyethylene, Merlon, high density are gathered Ethylene, politef ptfe or teflon (r), glass, quartz, metal (such as aluminum), transparent material, polysilicon etc..Accumulation Device hole top 1109 and 1110 also comprises to adjust screw, its removable with from accumulator room 1115 and 1116 introduce or remove gas or Liquid.Preferably, valve 1112 and 1111 can be started to discharge Fluid pressure in accumulator room 1115 and 1116, otherwise Fluid pressure Then retain wherein.Help microfluidic device is properly placed in other instruments using notch 1117, for example, for operation or Analysis microfluidic device or the instrument of the reaction wherein carrying out.

Fig. 3 is the simplified flowchart showing the operation method of microfluidic device according to an embodiment of the invention.? In embodiment shown in Fig. 3, there is provided 48 panel layouts of digital array.The is applied to pressure source 102 (receiving accumulator) One pressure, which results in starting of lock-in control line 110 (310).In a specific embodiment, described first pressure is 30psi.In other embodiments, other pressure are adopted according to concrete application.Referring to Fig. 1, all valves in array 106 (not shown) can be closed due to the pressure of section 114.Pressure is applied to section 116 by check valve 112, and this can lead to valve The closing of 118a-118e and locking.The overhead portion of section 116 can stop mensure input line from being closed due to starting of section 116 Close.Close and close the input flowing through manifold 135 for the fluid in the pass of valve 118, thus 6 mensure inputting 130 will be connected to mensure Input line is separated.

It should be noted that in the controlling stream shown in Fig. 3, not using pressure source 104.Therefore, do not have control line 120 start or The closing of separator valve 124.Therefore, respond to starting of lock-in control line 110, accommodate valve 128 and close and locking.Therefore, logical After crossing receiving accumulator applying first pressure (for example, 30psi), described array valve cuts out and valve 118 and 128 cuts out and closes Lock.

The first pressure (312) reducing pressure source 102 makes array valve (not shown) reopen.In some embodiments In, process (312) removes pressure so that pressure applied is zero.As noted above, valve 118 and 128 can cut out State keeps locking.Be supplied from sample input or mouth 140 sample be loaded into by sample input line 142 anti-in (314) array Answer room.The design of chip provides by sample input line 142, measures input line 132 or both loadings.Because here embodiment party In case, measure input line and closed by locking-valve 128, load and carry out through sample input line.After completing sample loading, to pressure Power source 102 applies second pressure (316) again, thus closing closed array valve and separating the sample in described reative cell.In some enforcements In scheme, second pressure is equal to first pressure (for example, 30psi), although this is optional in the present invention.Other are suitable to close The pressure of closed array valve is within the scope of the present invention.Although additionally, removing first pressure in some embodiments, this Optional in the present invention, because reduced pressure can still be provided by some embodiments to nonzero value loads filling of sample Sufficient flow of fluid.

Therefore, the embodiment that Fig. 3 shows provide 48 samples loading (that is, the sample input line shown in 24 × Device both sides).As discussed below, programmable digital array as herein described also provides other sample layout.

It should be understood that the concrete steps shown in Fig. 3 provide one kind of the microfluidic device of operation one embodiment of the invention Ad hoc approach.Other sequence of steps are implemented also dependent on some alternate embodiments.For example, some replaceabilities of the present invention are real The scheme of applying can implement step listed above in a different order.Additionally, single step shown in Fig. 3 can comprise multiple sub-steps Suddenly, it can be implemented with multiple orders according to the requirement of single step.Additionally, being added according to specific application or removing other Step.Those of ordinary skill in the art are it will be appreciated that many change, improve and replace.

Fig. 4 is a kind of simple flow of operation of display method of microfluidic device according to another embodiment of the invention Figure.Different from 48 panel layouts described by Fig. 3, provide single sided board layout in the embodiment depicted in fig. 4.To pressure source 104 apply first pressure (410), and this leads to starting of lock-in control line 120.Pressure can apply to close by check valve 127 With the valve 126 in locking section 125.Therefore, do not have the flowing through sample input line 142 in this layout.Elevated bridge section 122 can transmit control pressure to close separator valve 124.

Control pressure in lock-in control line 120 can be transferred to related to 5 in 6 mensure input lines 132 by section 123 The valve 134 of connection.Therefore, valve 134 will be closed and only passes through the mensure input line of low order end so that flowing.

Mensure input line through low order end loads single sample (412).Section 116 here due to lock-in control line 110 It does not actuated in layout, manifold 135 is opened, in so that low order end sample is inputted, contained single sample is provided to whole samples Input line 132.Therefore, input the described array of loading after measured using the single sample inputting from low order end sample determination.Right Acute build up of pressure device 102 applies second pressure, starts lock-in control line 110 and closes the array valve being connected with section 114.Therefore, Using different controlling streams, the multiple layouts using single programmable digital array are possible, for example, single sided board layout.

It should be understood that the concrete steps shown in Fig. 4 provide the one of microfluidic device according to another embodiment of the invention Plant particular methods of operation.Other sequence of steps are implemented also dependent on alternate embodiment.For example, some replaceabilities of the present invention Embodiment can implement step listed above in a different order.Additionally, single step shown in Fig. 4 can comprise multiple sub-steps Suddenly, it can be implemented with multiple orders according to the needs of single step.Additionally, being added according to specific application or removing other Step.Those of ordinary skill in the art are it will be appreciated that many change, improve and replace.

Fig. 5 is the simple flow of a kind of method showing operation according to the microfluidic device of further embodiment of the present invention Figure.In the embodiment depicted in fig. 5, there is provided 12 panel layouts of digital array.First pressure is applied to pressure source 104 (510).In a specific embodiment, described first pressure is 45psi, although this is optional in the present invention, The pressure being suitable for using other.As the discussion of the process (410) about Fig. 4, start lock-in control line 120.By check valve 127 applying pressure with close and locking section 125 in valve 126.Therefore, do not have in this layout through sample input line 142 flowing.Elevated bridge section 122 can transmit described control pressure to close separator valve 124.As discussed below, separator valve Temporarily Closed can stop accommodate valve 128 subsequent control line 110 start under effect close.Section 123 can be by locking control The control pressure of line 120 processed is transferred to measure 5 valves being associated 134 in input line 132 with 6.Therefore, valve 134 can be temporary When close.

Second pressure (512) is applied to pressure source 102, starts lock-in control line 110.In a specific embodiment In, described second pressure is 30psi, less than first pressure (for example, 45psi).As discussed below, described second pressure foot It is not turned on the separator valve 124 closed in process (510) to close predetermined valve.Other can be used to press according to concrete application Power.Section 114 start the valve that can close in closed array 106.Pressure can apply to section 116 through check valve 112, and this can lead to valve The closing of 118a-118e and locking.The overhead portion of section 116 can stop mensure input line from being closed due to starting of section 116 Close.Close and close the input flowing through manifold 135 for the fluid in the pass of valve 118, thus 130 6 mensure being connected will be inputted with measuring Input line each separate.

Initial from control line 102 starts different, and separator valve cuts out by initial the starting of control line 104, thus providing Starting of programmable sequences, wherein control line 102 does not lead to and measures valve 128 locking that input line 132 is associated.This multiple Control characteristic provides the asynchronous logic function using some embodiments of the invention, will not be obtained using conventional microfluidic device Obtain this characteristic.

It is applied to the reduced pressure of pressure source 102 to such as zero (514).Because section 116 is in check valve 112 downstream, After section 116 revocation is started, valve 118 continues to be latched in closed position, and this stops 6 flow of fluids measuring between input line. The valve opened in described array is started in the revocation of section 114, makes to load sample in subsequent loading process.As discussed above , valve 128 is non-latching in process (512).

It is applied to the reduced pressure of pressure source 104 to such as zero (516).Again opened by reducing the pressure of section 123 Open valve 134.As discussed above, provide input using mensure input 130 for the reative cell in array.By reducing section Pressure in elevated bridge section 122 in 121 is reopening separator valve 124.Because this stage in programming process is to control line 110 Do not apply pressure, so valve 128 is maintained at opening.Although shown is that pressure drop is as low as zero, these tools Body pressure is not essential to the invention, the pressure being suitable for using other.

Measure from 6 and input, be loaded into sample (518).By going of the manifold 135 that led to due to the locking of valve 118 before Functionalization keeps measuring the separation between input.Therefore, in this layout, there is provided (6 measure input × dress to 12 panel layouts 2 sides put).Pressure source 102 is applied with the 3rd pressure (520) to close the array valve being associated with section 114.Further, since Separator valve 124 is switched on again, so valve 128 is blocked cut out.

It should be understood that the concrete steps shown in Fig. 5 provide the one of the microfluidic device operating further embodiment of the present invention Plant concrete grammar.Other sequence of steps are implemented also dependent on alternate embodiment.For example, the alternate embodiment of the present invention Step listed above can be implemented in a different order.Additionally, single step shown in Fig. 5 can comprise many sub-steps, it can Implemented with multiple orders according to single step.Additionally, being added according to specific application or removing other step.This area is general Logical technical staff is it will be appreciated that many change, improve and replace.

Table 1 provides opening of one of the multiple possible panel layout of programmable digital array microfluidic device as herein described Dynamic pressure and loading process.The pressure exemplary pressure that pressure source 102 and 104 is applied illustrates, although other are suitable for Pressure be included in the scope of embodiment of the present invention.

Table 1

Programming step	48 panel layouts	1 panel layout	12 panel layouts
				1	102=30 psi	104=30 psi	104=45 psi
2	102=0 psi	It is loaded into sample	102=30 psi

3	It is loaded into sample	102=30 psi	102=0 psi
				4	102=30 psi	104=0 psi
5			It is loaded into sample
				6		102=30 psi

The integration of check valve and separator valve and digital array provides multiplex using single high-density digital array chip Family programs layout.Therefore, some embodiments of the present invention provide the saving of cost and the raising of experiment flux.Described herein Serial or asynchronous logic provide enhanced apparatus function, and this is because the state of particular valve not only depends on and particular valve fluid Starting of the control line of connection, additionally depends on the state of the separator valve that independent control line is started.Therefore, the order of pressurization is determined Determine the logic output of panel layout.Therefore, provide can not using conventional microfluidic body device for some embodiments of the present invention The device layout obtaining and function.

According to one embodiment of the invention, there is provided the microfluidic device with multiple control lines is configured Method.Serial or asynchronous logic are implemented according to the method.Methods described includes starting the first control line and valve is placed in One state.Valve (can be one of multiple valves) can respond to described first starting of control line and be placed in off.For example, Separator valve 124 can respond to the pressurization applying pressure and control line 120 of interface accumulator 104 and cut out.As in this explanation Book in full in be described more fully and described more particularly below, due to separator valve be placed in valve 128 and accommodate accumulator 102 it Between, separator valve can provide multiple control function, and this provides serial logic operation.

Methods described also includes, and after valve is placed in first state, starts and is operable to for valve group to be placed in second Second control line of state.The situation that valve is in first state (cutting out) stops valve group from being placed in (for example, the closing of the second state State).Valve 128 is stoped to respond to opening of control line 110 as shown in figure 1, closing separator valve 124 before starting control line 110 Move and close (and locking).Therefore, some embodiments of the present invention provide serial logic function, the order that wherein valve starts Create the different outcomes selected from several end-state.In this example, the initial of control line 110 starts the pass leading to valve 128 Close and locking.On the other hand, the initial of control line 120 starts the closing leading to separator valve 124, when keep in off position when its Stop closing and the locking of valve 128.While figure 1 show that applying, to independent pressure source 102 and 104, the control that pressure is started Line 110 and 120, but other start method and comprise within the scope of the present invention.

According to another embodiment of the invention, there is provided execute the another kind side of serial logic using microfluidic device Method.The method that the microfluidic device with multiple control lines is configured includes setting up microfluid by starting the first control line The first state of device, then starts the second control line.The first state of microfluidic device may include and responds to a plurality of control line Start valve be set on or closed mode.For example, as described above, starting control line 120 before starting control line 110 Lead to the closing of separator valve 124.Therefore, valve 128 does not respond to starting of control line 110 and locking is closed.

Methods described also includes setting up the second state of microfluidic device by starting the second control line, then starts described First control line.The programmable features of microfluidic device as herein described provide the microfluidic device starting order depending on valve Different end-state.Therefore, in this example, start the pass that control line 110 leads to valve 128 before starting control line 120 Close the closing with locking and separator valve 124 thereafter.It is adiaphorous for closing separator valve after locking-valve 128, because check-valves 112 constant pressures keeping the control line to check valve downstream, stop the closing of separator valve from affecting the state of microfluidic device.

Implement the example of serial logic as microfluidic device, the first state of microfluidic device can comprise to close or block The first input line group (input line 142 that for example valve 126 is blocked) of state and with multiple reative cells in described microfluidic device The the second input line group (such as input line 132) being in fluid communication.Described second state can comprise many with described microfluidic device The described first input line group (such as input line 142) that individual reative cell is in fluid communication, and close or blocked state second defeated Enter line group (input line 132 that for example valve 128 is blocked).Second state also can comprise valve group (the such as valve 118a- of closed mode 118e), thus stoping fluid through connecting the manifold flow of the second input line group.

It is noted that while separator valve is started by control line 120, other embodiments can be opened by control line 110 using another kind Dynamic separator valve, using as the replacement of shown separator valve or supplement.Thus, for example, overhead route can lead to from accommodating accumulator 102 To the extra separation valve between interface accumulator 104 and check-valves 127.To the control connecting to interface accumulator 104 The section 125 of line is closed this extra separator valve before applying pressure and valve 126 can be stoped to respond to starting and closing of control line 120 And locking.Therefore, while figure 1 show that being embodied as, the invention is not restricted to this and being embodied as, other microfluidic device design packet Containing within the scope of the invention.

Although additionally, the embodiment shown in Fig. 1 and replaceability geometric shape discussed above employ by two A row replacement plus-pressure in control line and the separator valve being started and two levels providing multiple control, the reality of the present invention The scheme of applying provides other levels of multiple control.For example, can by other separate valves (for example, electrostatic starts) integration in a subtle way In the design of fluid means, to provide the control of the 3rd level.In addition to the starting of control line shown in except 2, other to this The initial logic output starting meeting driving means state of separate valves.

Fig. 7 shows the method that the programmable microfluidic device to one embodiment of the invention is programmed.In Fig. 7 institute In the embodiment shown, described microfluidic device is programmed for by the reative cell of described microfluidic device be separated into 48 independent Panel.The specific number of panel depends on the actual design of microfluidic device, and can be for not in the embodiment of replaceability Same number, such as 12 or 192 panels.The reative cell of each separate panels can fill different samples, in the present embodiment Provide detection while up to 48 different samples.

As shown in fig. 1, microfluidic device has the reaction site being arranged in array 106 or reative cell array.Described Reaction site is in fluid communication with the first input line group 132 and the second input line group 142.Methods described includes starting (710) can quilt Operation is in order to block the first valve group of (712) first input line groups.For example, the first valve group is valve 128, also referred to as can be used by operation To stop or to close the receiving valve of input fluid line 132.For example by accommodate accumulator 102 apply pressure closes valve 128 Fluid is stoped to flow by input line 132.Because unidirectional or check-valves 112 presence make control line 110 become lock-in control line, After accommodating valve 128 closing, they remain turned-off.

Methods described also includes starting the second valve group that (714) are operable to separation (716) described reaction site.As Shown in Fig. 6 b, reative cell 630 is limited in the elastomer layer 603 of microfluidic device.Described second valve group is arranged in array, And it is shown as valve 615 in Fig. 6 a.In addition to the first valve group 128 and the second valve group 615, the 3rd valve group 118a-118e and described the One valve group is started simultaneously.It is locking after closing that the presence of check-valves 112 also makes valve 118a-118e.Embodiment party shown in Fig. 1 In case, described first valve group and described second valve group respond to the pressure that control line 110 is applied and start simultaneously or close.

After described first, second, and third valve group is closed, described second valve group revocation starts (718), for example, by fall The pressure that low or elimination is applied to control line 110 by pressure source 102.The unlatching leading to described valve is started in the revocation of described valve, this Allow fluid flow the input line being connected with valve.Multiple samples are loaded into reaction site (720) by the second input line group 142.In Fig. 1 In, sample is included in sample input or mouth 140.The both sides of described microfluidic device respectively comprise 24 different input ports, carry Totally 48 input ports are supplied, although this concrete numeral can change according to the specific design of microfluidic device.Sample port can comprise 48 Individual different sample or when providing single sample in multiple input ports, can comprise the less sample of sum.Therefore, although table 1 is related to 48 panel layouts, and programmable microfluidic device provides up to 48 indeed according to the sample providing in 48 sample ports Independent panel.After sample is loaded into reaction site, start the second valve group for the second time so that reative cell is separated (722).

It should be understood that the concrete steps shown in Fig. 7 provide of the microfluidic device programming to one embodiment of the invention Plant concrete grammar.Other sequence of steps also dependent on alternate embodiment implement.For example, the replaceability embodiment party of the present invention Case can implement step listed above in a different order.Additionally, single step shown in Fig. 7 can comprise many sub-steps, its Can be implemented with multiple orders according to single step.Additionally, being added according to specific application or removing other step.This area Those of ordinary skill is it will be appreciated that many change, improve and replace.

Fig. 8 shows the method that the programmable microfluidic device to another embodiment of the invention is programmed.Fig. 8 shows The method shown provides and the programmable microfluidic device having a single panel and all reative cells comprise same sample is carried out The method of operation.As shown in figure 1, described programmable microfluidic device has and the first input line group and the second input line group fluid The reaction site array of connection.In exemplary shown in Fig. 8, described first input line group is and sample input The input fluid line 142 of 140 connections, described second input line group is and measures the input fluid line 132 that input 130 is connected.Will First input line group is started for closed position (810), the flowing (closing) (812) through the first input line group for the obstruction.Therefore, exist In one example, interface accumulator 104 pressurization is led to the closing of valve 126, stops fluid from flowing through input line 142.Check-valves 127 presence leads to the first valve group to be latched in closed position.

Methods described also includes starting first that (814) are operable to block (816) second input line group subgroups The second valve group divided.Referring to Fig. 1, the Part I of described second input line group is the part of the input line 132 under valve 134, Described second valve group is valve 134, and operable its is measured in input 130 6 related input lines 132 to 6 with blocking or closing 5.Therefore, in the present embodiment, the subgroup of input line is 5 in 6 input lines, using the low order end shown in Fig. 1 Mensure input port (being connected with the input line of low order end) sample being loaded into reative cell or site is provided.It is not included in subgroup Input line (i.e. the input line of low order end) is opened and is reacted for being loaded into sample as described more fully below Room.Do not measure input port using other in this is embodied as.In the embodiment depicted in fig. 1, interface accumulator 104 applies Pressure start valve 126 and valve 134 simultaneously.

Referring to Fig. 1, the starting of the valve that is not carried out carrying out by the use of control line 110.Therefore, sample is loaded into low order end Measure input port sample can be led to flow through section 116 and flow to the Part II of described second input line group by manifold 135 (i.e. the part of the input line 132 on valve 134).Sample is loaded into the reaction in array 106 by described second input line group In room or site (818).It should be noted that being made by the starting of section 116 through uninflated control line 110 in the present embodiment Obtain manifold and disable (i.e. the flowing through manifold is prevented from).Therefore, manifold open and input line 132 Part II rise Beginning section connects all 6 input fluid lines.Through manifold 135, sample can be allocated to the component of input line 132, Flow through 24 input lines eventually, through the valve 128 opened.Therefore, single sample is supplied to all reative cells in array 106. In order to separate the sample in reative cell (822), start the 3rd valve group (820), such as the receiving valve 615 shown in Fig. 6 a.By making The pressure closing the valve 615 in closed array 106 can be provided with accommodating accumulator 102 to control line 110 applying pressure.

It should be understood that the concrete steps shown in Fig. 8 provide the microfluidic device to another embodiment of the invention compiling A kind of concrete grammar of journey.Other sequence of steps are implemented also dependent on alternate embodiment.For example, the replaceability of the present invention Embodiment can implement step listed above in a different order.Additionally, single step shown in Fig. 8 can comprise multiple sub-steps Suddenly, it can be implemented with multiple orders according to single step.Additionally, being added according to specific application or removing other step. Those of ordinary skill in the art are it will be appreciated that many change, improve and replace.

Fig. 9 shows the method for simplifying being programmed of the microfluidic device to further embodiment of the present invention.Described Programmable microfluidic device has and the first input line group, the second input line group and the manifold stream being connected described second input line group The reaction site array of body connection.Methods described includes starting (910) and is operable to block (912) first input line groups First valve group.Referring to Fig. 1, the first valve group can be the valve 126 in check-valves 127 downstream.The presence of check-valves 127 makes valve 126 open It is latched in closed position after dynamic.Start the second valve group (914) to block the Part I (916) of the second input line group subgroup.Right Control line 120 applies to lead to starting of valve 126 and valve 134 from the pressure of interface accumulator 104.

Referring to Fig. 1, the Part I of described second input line group is input line 132 part under valve 134, and the second valve group is Valve 134, it is operable to block or closes 5 being measured in 6 input lines that input 130 is connected with 6.Therefore, exist In the present embodiment, the subgroup of input line is 5 in 6 input lines.Because valve 134 is not by unidirectional valve blocking, when again opening When putting these valves, the Part I that fluid can flow through all 6 input lines 132 flows to manifold 135.Embodiment party shown in Fig. 1 In case, apply the pressure from interface accumulator 104 and start valve 126 and valve 132 simultaneously.

Methods described also includes starting the 3rd valve group that (918) are operable to make manifold disable (920).Show at one In example property embodiment, starting of the 3rd valve group (for example, accommodates accumulator including to the pressure source being in fluid communication with the second valve group 102) apply predetermined pressure.Referring to Fig. 1, valve 118a-118e is started by the section 116 applying pressure to control line 110 The flowing through manifold can be blocked, to provide the separation to the fluid flowing through input line 132.The presence of check-valves 112 can lead to valve 118a-118e is latched in closed position.As described more completely below, the deactivation of manifold 135 makes 12 sample (microfluidic devices Every side 6 sample) input port 130 loading reative cell can be measured from 6.As discussed in the embodiment of Fig. 7, by inciting somebody to action Identical sample is supplied to more than one mensure input and can be loaded into less than 12 samples.It will be appreciated by those skilled in the art that it is many Change, improve and replace.

Referring back to Fig. 1 it should be noted that before starting control line 110 initial the starting of control line 120 do not only result in valve 126 He The closing of valve 134 and locking, also result in the closing of separator valve 124.Because separator valve is arranged in receiving accumulator 102 and valve 128 Between, the closing of separator valve can stop valve 128 from responding to control line 110 to start and close.In the embodiment depicted in fig. 9, Pressure for starting control line 120 is predetermined pressure, and it be enough to stop starting of the control line 110 leading to valve 128 to be closed. As shown in table 1, the pressure of 45psi be enough to, when control line 110 is applied with the pressure of 30psi, valve 128 is maintained at open position. Specific value 45psi and 30psi are not essential to the invention, can implement this multiple control function using other pressure.

Methods described also includes revocation and starts the second valve group (922).Generally, by applying to the first of first pressure source Reduced pressure for example to cancel to 0psi starts the second valve group, so that it takes open position.It is applied to the pressure of control line Reduce the position making the flexible membrane return being deflected into described input fluid line substantially not deflect, flow through the fluid of input fluid line Flowing recovers.After reopening valve 134, the sample providing in measuring input 130 is written into reaction site (924).As Fig. 1 Shown, the second input line group Part I (part of the input line 134 for example under valve 134) of each with measure input port It is in fluid communication, described mensure input port is arranged to receive one of multiple samples.Sample flows through the first He of the second input line group Part II enters reaction site.As shown in figure 6 a and 6b, described reaction site can be formed in the form of multiple reative cells, wherein Each reative cell is passed through through hole and is in fluid communication with the Part II of input fluid line 132, and described through hole passes through input fluid line to arrive Reach described reative cell, described reative cell can be disposed in the elastomer microfluidic device layers on the layer containing fluid line.

As shown in figure 1, the element branches of input line 132 being located on valve 134 become four input line groups, itself and array Reaction site in 106 or room are in fluid communication.6 different samples are provided by measuring at each in input port, can be in an array Definition 12 panels (because Fig. 1 illustrate only the first side of microfluidic device) altogether.As aobvious and easy to those skilled in the art See, can lead to produce less than 12 panels using a sample in more than one input port.In order to separate reaction site, Start the 4th valve group (926), such as valve 615.

In some implementations, methods described also includes starting valve to stop the closing of the 5th valve group, and the wherein operable 5th Valve group is to close the Part II of the second input line group.As shown in figure 1, the closing of separator valve 124 stops valve 128 from responding to control The starting and close of line 110.In the embodiment depicted in fig. 9, the pressure of the 45psi being applied to control line 120 when initial is led Cause the closing of separator valve, make the Part II of input line 132 keep it turned on manifold simultaneously and stopped by the closing of valve 118a-118e With.Separator valve can be started by applying pressure to control line or can otherwise start, including but not limited to mechanical, quiet Electricity etc..Using in the application based on the control line of pressure, multiple valves and/or valve group can be started using single control line simultaneously. Therefore, when pressurizeing to interface accumulator 104, separator valve 124 and valve 126 and valve 134 respond to opening of control line 120 simultaneously Move and actuated.

The programmable nature of microfluidic device provided herein is proved by following facts: changes the order meeting that valve starts step Lead to the valve in array to be in different states, and thus make array be in different states.For example, the enforcement shown in Fig. 9 In scheme, by following steps, 12 samples can be loaded into array, described step is: close separator valve 124, then to control line 110 applying pressure, this makes manifold 135 disable, then reduces the pressure to accumulator 102, then reduces the pressure to accumulator 104 Power.In this example, pass through to keep when accumulator 102 applies stoping valve 128 from responding to the pressure of accumulator 104 in pressure The closing started of the control line 110 being caused using the pressure of accumulator 102 applying.On the other hand, by accumulator 102 Apply release during pressure and be applied to the pressure of accumulator 104 to lead to the closing of valve 128.Therefore, by change valve start suitable Sequence achieves two different conditions of microfluidic device, is loaded into sample or the ability of closing input line 132 by input line 132. This programmable features provides using the not obtainable benefit of conventional design.

It should be understood that the concrete steps shown in Fig. 9 provide the one of the microfluidic device programming further embodiment of the present invention Plant concrete grammar.Other sequence of steps are implemented also dependent on some alternate embodiments.For example, some replacements of the present invention Property embodiment can implement step listed above in a different order.Additionally, single step shown in Fig. 9 can comprise many height Step, it can be implemented with multiple orders according to single step.Additionally, being added according to specific application or removing other step Suddenly.Those of ordinary skill in the art are it will be appreciated that many change, improve and replace.

It should be understood that the invention is not restricted to concrete grammar as herein described, scheme and reagent etc., those skilled in the art can manage Solve these change.It will also be understood that term used herein is intended merely to for describing specific embodiment it is no intended to limit The scope of the present invention processed.It should also be noted that as used herein and in claims, unless in addition clear and definite in context Point out, do not indicate and during quantity, include plural form.Thus, for example, referring to one or more cells when referring to " cell ", this Skilled person's its equivalent known.

Except as otherwise noted, all scientific and technical terminologies used herein have the technical field of the invention those of ordinary skill The same meaning being generally understood that.The details of embodiment of the present invention and its various features and advantage is with reference in accompanying drawing and following theory Bright book detailed description and/or the non-limiting embodiments of explaination and embodiment are explained more fully.It should be noted that in accompanying drawing Shown feature is not necessarily to scale, and as understood by a person skilled in the art, even if herein not Explicitly point out, the feature in an embodiment can be used for other embodiments.Description to known assemblies and treatment technology can It is omitted to avoid and embodiment of the present invention is produced with unnecessary obscuring.Example used herein is only to facilitate understand this The mode that invention can be carried out, and make those skilled in the art implement embodiment of the present invention further.Therefore, the reality of this paper Example and embodiment are understood not to limit the scope of the present invention, and the scope of the invention is only by claims and applicable law Limit.Furthermore, it should be noted that similar reference in several views of accompanying drawing represents similar part.

Therefore, following present " definition " part, wherein in order to clearly specifically define some arts related to the present invention Language, but it is defined consistent to the understanding of these terms with those skilled in the art.Describe specific method, equipment and material Material, although any method similar or equivalent with those described herein and material can be used for implementing or test the present invention.This All lists of references that literary composition is related to pass through overall being incorporated herein by reference.

Definition

Pna is peptide nucleic acid(PNA) (peptide nucleic acid)

Lna is lock nucleic acid (locked nucleic acid)

Da is dynamic array (dynamic array)

Pcr is polymerase chain reaction (polymerase chain reaction)

Bsa is bovine serum albumin (bovine serum albumin)

Fret is FRET (fluorescence resonance energy transfer) (fluorescence resonance energy transfer)

Gt is gene type (genotyping)

Peg is Polyethylene Glycol (polyethylene glycol)

Plp is padlock probe (padlock probe)

Terms used herein " neighbouring " typically refers to the position that primer is with respect to probe on target nucleic acid analyte complementary strand.Draw Thing and probe can be spaced about 1 to about 20 nucleotide, more specifically, about 1 to about 10 nucleotide, or can be immediately adjacent to one another.

Terms used herein " analyte " typically refers to the core defined below of the inventive method detection to be used or quantitation Acid molecule or nucleic acid molecules mixture.Term " target nucleic acid analyte " and " nucleic acid analyte " can " be divided with term in the present invention Analysis thing " used interchangeably.

Terms used herein " complementary " or " complementary " can be included in polynucleotide under the salt of permission and temperature conditionss and pass through The natural combination of base pairing.For example, sequence " a-g-t " combines its complementary series " t-c-a ".Mutual between two single chain molecules Benefit can be " part ", only few of which nucleic acid combine, or the complementation between two single chain molecules single chain molecule it Between be completely when having overall complementary.Complementarity between nucleic acid chains has to the hybridization efficiency between nucleic acid chains and intensity Important function.This is even more important in the amplified reaction of the combination depending on nucleic acid interchain and the design and use of molecule.

Terms used herein " covalent attachment " typically refers to a molecular structure and passes through covalent chemical bond and another molecule The connection of structure.

Terms used herein " dyestuff " typically refers to absorption any of electromagnetic radiation when wavelength is more than or equal to 340nm to be had Machine or inorganic molecule.

Terms used herein " fluorescent dye " typically refers to by electromagnetic radiation source (for example, lamp, photodiode or sharp Light) irradiate after sent by Fluorescence Mechanism longer wavelength electromagnetic radiation any dyestuff.

Terms used herein " gt sample buffer " typically refers to can be by the bound site of reaction channel and chamber surface in da chip Select the buffer of closing.Described buffer prevents the loss of reactive component when chip is loaded into and processes or react.For reagent cost For, it also can be by other taq-gold polymerase using reduction less than about 80%.20 × gt buffer may include following Combination: glycine betaine (fw 117.15), bsa,T20 (in pbs) (thermo scientific, Rockford, il),(in pbs) (thermo scientific, rockford, il),(in tbs) (thermo scientific, rockford, il),T20 ( In tbs) (thermo scientific, rockford, il), glycerol, peg 20,000, peg mme550, peg mme5000 With tween 20.

Terms used herein " homogeneity measures (homogenous assay) " typically refers to detection or quantitative nucleic acid analyte Method, it need not measure post processing to record measurement result.Homogeneity measures and can carry out in the pipe of closing or microfluidic arrays, And do not need further to add reagent after mensure is initial or supplement chemicals to record result.Homogeneity measures can real time record The result measuring is it means that measurement result continuously can be recorded when measuring and being in progress in time.

Terms used herein " hydrolysis probes ", in United States Patent (USP) no.5, has carried out general description in 210,015, and it passes through Entirety is incorporated herein by reference.Hydrolysis probes make use of 5 ' present in heat stability taq polymerase used in pcr reaction- Nuclease (Probe technique, applied biosystems, foster city ca).Use fluoroscopic examination Dyestuff (as fluorescein) and receive dyestuff or substance markers hydrolysis probes are quenched.Generally, fluorescent dye is covalently attached to the 5 ' of probe End, quencher is connected to 3 ' groups of probe, and when probe is complete, the fluorescence of detection dyestuff is due to FRET (fluorescence resonance energy transfer) (fret) it is quenched.Probe can be in the anneals downstream of a primer, and described primer limits pcr reaction nucleic acid target analyte amplification One end of target position.Using the polymerase activity of taq enzyme, the amplification of target nucleic acid analyte by probe upstream a primer and The the second primer guiding annealed positioned at probe downstream but on the relative chain of target nucleic acid.With the extension of forward primer, taq is polymerized Enzyme reaches the region of label probe annealing, and probe-template crossbred is identified as substrate, and the phosphodiester bond of hydrolysis probes. Hydrolysis irreversibly discharge the quenching effect to reporting dyes for the quencher dyes, thus result in and follow with each continuous pcr Ring, detector fluorescence increases.Specifically, the hydrolysis probes of the present invention can in common 8-mer in people and other transcript profile or 9-mer motif simultaneously can be by having about 70 DEG C of high t using lna analog_m.

Terms used herein " labelling " refer to can be used for provide can detect and/or can quantifiable signal any atom or point Son.Specifically, described labelling may be connected to nucleic acid or protein.Labelling can provide and can pass through fluorescence, radioactivity, colorimetric, x The signal of the detection such as optical diffraction or absorption, magnetic, enzymatic activity.

Terms used herein " nucleic acid " typically refers to cdna, dna, rna, its single-stranded or double-stranded and any chemical modification, such as Pna and lna.Lna in United States Patent (USP) no.6,794,499,6,670,461,6,262,490 and 6, described in 770,748, and lead to Cross to quote in full and be expressly incorporated herein.Nucleic acid can be any size.Nucleic acid is modified and be may include interpolation chemical group, and it is to single nucleic acid Base or nucleic acid integrally add extra electric charge, polarity, hydrogen bond, electrostatic interaction and function.This modification may include modification Base such as 2 '-position is sugar-modified, the modification of 5- position pyrimidine, the modification of 8- position purine, the modification of cytosine exocyclic amine, 5-bromouracil Replacement, backbone modification, methylate, the different cytidine of unusual base combinations of pairs such as isobase and isoguanidine etc..Nucleic acid may be from Full chemistry building-up process (as the chemosynthesis of solid phase mediation) or come from biogenetic derivation (as passed through from nucleic acid can be provided almost Separate in any species) or come from process (such as dna duplication, the pcr expansion being related to operate nucleic acid by biology tool Increasing, reverse transcription) or the combination from these processes.

Terms used herein " nucleic probe " is to carry the nucleic acid of at least one covalent attachment dyestuff (as fluorescent dye).Tool For body, probe do not comprise with for causing the complementary sequence of the sequence that pcr reacts.

Terms used herein " padlock probe " or " plp " typically refer to the linearly few core with about 100 base pairs length Thuja acid.The sequence of plp 3 ' and 5 ' end is complementary with the neighbouring sequence in target nucleic acid analyte.Central incomplementarity region in plp There is " sequence label " that can be used for identifying specific plp.The flank of described sequence label can for universal primer site or unique and/ Or specific primer site, it allows pcr to expand described sequence label.After being hybridized to target, the 5 ' of plp and 3 ' ends are tight Adjacent and subsequently can be connected.Products therefrom is the cycling probe molecule connecting target nucleic acid analyte.For example can usePcr expands to the label area of cyclisation plp and quantitative and/or detection in real time.The presence of amplicon and amount can Presence with target sequence in sample and amount are associated.The description of plp be see, for example, with landegren etc., 2003, padlock And proximity probes in situ and array-based analyses:tools for the post- Genomic era, comparative and functional genomics 4:525-30；Nilsson etc., 2006, Analyzing genes using closing and replicating circles trends biotechnol.24: 83-8；Nilsson etc., 1994, padlock probes:circularizing oligonucleotides for Localized dna detection, science 265:2085-8.Above-mentioned list of references passes through to quote in full to be expressly incorporated herein.

Terms used herein " pcr " typically refer to for expand, detect or quantifying analytes specific region method.Ability Field technique personnel will understand that has several changes in basic pcr technical foundation, such as allele specific pcr, set pcr (assembly pcr) or polymerase cycle set (polymerase cycling assembly, pca), colony pcr, untwist Specificity (intersequence-specific, issr) pcr between enzyme dependent amplification, thermal starting pcr, sequence, reverse pcr, Connect mediation pcr, methylation-specific pcr, many sites join dependency probe amplification, many sites pcr, nido pcr, overlap extension Pcr, quantitative pcr, quantitative pcr in real time, rt-pcr, hot asymmetric interlaced (thermal asymmetric interlaces, Tail) pcr, landing pcr and pan-ac.How in addition, those skilled in the art will appreciate that these change application basic Pcr is technical.

Terms used herein " purification " typically refers to protein, polypeptide according to electric charge, molecular size or binding affinity Or nucleic acid any process detached with other compositions or compound.

Terms used herein " quencher " typically refers to reduce the dyestuff of the fluorescent emission of another dyestuff.

Terms used herein " inquiry (querying) " typically refer to determine target specific probe whether with nucleic acid analyte Related (for example, connect or link), and optionally in quantitative sample target specific probe amount.

" sample " used herein typically refers to the tissue from human or animal or fluid sample, including but not limited to blood plasma, Serum, spinal fluid, lymph fluid, the outer section of skin, respiratory system, intestinal and urogenital tract, tear, saliva, hemocyte, swell Tumor, organ, the sample of tissue and Cell culture invitro component.Specifically, sample can be unicellular, paraffin-embedded tissue sample Product and acupuncture biopsies.Additionally, sample may include environmental sample, such as lake water and foodstuff samples.

Word " basic purification " used herein or " being basically separated " generally include and obtain, separate from their natural surroundingses Separate nucleic acid or aminoacid sequence, they at least do not contain about 60%, especially at least about 75%, most particularly at least about 90% other components associated there, and include restructuring or the separate nucleic acid thing cloned and chemosynthesis analog or be Unite biosynthetic analog.

In view of the very big multiformity of polymer chemistry, precursor, synthetic method, reaction condition and available additive, Substantial amounts of useful elastomers system is had to can be used for manufacturing elastic module, layer, film, miniature valve, pump etc..The change of material therefor is one Can be due to needing specific material behavior in the case of a little, that is, solvent resistance, rigidity, breathability or temperature stability are determining.Have Very eurypalynous elastomeric polymer.Even if providing being briefly described it is therefore an objective to show to adopt of modal elastomer type herein The polymer of relative " standard ", there is also multiple possibility of combination.Conventional elastomeric polymer includes polyisoprene, polybutadiene Alkene, polychlorobutadiene, polyisobutylene, poly- (s-B-S), polyurethane and silicone or polysiloxanes.

Polyisoprene, polybutadiene and polychlorobutadiene are polymerized by diene monomers, therefore polymerization when each Monomer has a double bond.This double bond makes polymer pass through sulfuration, and (generally, sulfur is used for being formed between double bond by heating and hands over Connection) it is converted into elastomer.This makes to carry out the soft printing of easy homogenizing multilamellar by the incomplete vulcanization treating binder courses (homogeneous multilayer soft lithography)；Photoresist encapsulation can be carried out by similar mechanism (photoresist encapsulation).

Pure polyisobutylene does not have double bond, but is handed over by introducing the isoprene of a small amount of (about 1%) in polymerization Connection is thus be used as elastomer.This isoprene monomer provides pendency double bond on polyisobutylene skeleton, and it can be in subsequent as above institute Cure with stating.

Poly- (s-B-S) passes through active anionic polymerization, and (i.e. no natural chain termination walks in the reaction Suddenly) produce, therefore can there are " active " polymer ends in the polymerization species of solidification.This is the sky of photoresist package system So material standed for (wherein having unreacted monomer in a large number in the liquid layer being cast in cured layer top).Incomplete solidification makes The soft printing of homogeneous multilamellar (a is combined) can be carried out with a.This chemical property is additionally favorable for making to have an extra butadiene (" a ") And coupling agent, another layer (" b ") shortage butadiene (for the soft printing of homogeneous multilamellar).Sbs is " thermo-setting elastomer ", means Its fusing plastic (relative with elasticity) on uniform temperature；Reduce temperature and produce elastomer again.Therefore, layer can be tied by heating It is combined.

Polyurethane is made up of diisocyanate (a--a) and glycol or diamidogen (b--b)；Due to diisocyanate and glycol/ Amine has many kinds, the enormous amount of different types of polyurethane.However, the characteristic with respect to b for a of polymer makes them as rtv 615 are used for like that the soft printing of heterogenous multilayer: by one layer using excessive a-a, using excessive b-b in another layer.

Siloxane polymer has great structure change, and provides multiple commercial preparation.Discuss rtv's 615 Vinyl-to (si--h) crosslinked (the soft printing of heterogenous multilayer and photoresist encapsulation can be carried out), but this is only intended to siloxanes and gathers One of several cross-linking methods of compound chemistry.

Except using above-mentioned simply " pure " polymerization beyond the region of objective existence, cross-linking agent can be added.Some reagent are (as having for sulfuration The monomer of pendency double bond) it is suitable for the soft printing of homogenizing (a to a) multilamellar or photoresist encapsulation；In the method in two bullets Property layer in mix identical reagent.Complementary reagents (i.e. a kind of monomer has pendency double bond, and another kind has pendency si-h group) It is suitable for the soft printing of heterogeneous (a to b) multilamellar.Add mutual tonic to adjacent layer in the method.

It is below the non-exhaustive list of the elastomer being used in combination with the present invention: polyisoprene, polybutadiene, polychlorostyrene Butadiene, polyisobutylene, poly- (s-B-S), polyurethane and siloxane polymer；Or poly- (two (fluoroalkanes Epoxide) phosphine nitrile) (pnf, eypel-f), poly- (carborane-siloxanes) (dexsil), poly- (acrylonitrile-butadiene) (butyronitrile rubber Glue), poly- (1-butylene), poly- (chloro trifluoro ethylene-difluoroethylene) copolymer (kei-f), poly- (ethyl vinyl ether), poly- (two Fluorothene), poly- (difluoroethylene-hexafluoropropene) copolymer (viton), polrvinyl chloride elastic composition (pvc), polysulfones, poly- carbon Acid esters, polymethyl methacrylate (pmma) and politef (teflon).

Allcock etc., contemporary polymer chemistry, elastomer is described as being present in glass by the second edition Polymer at a temperature of between glass conversion temperature and condensing temperature.Elastomer display elastic characteristic is because polymer chain is easy Skeletal chain is made to untie under force in the motion that twists, the skeletal chain curling in powerless effect returns shape originally Shape.Generally, distortional elastomer when force is applied, reverts to original shape when removal force.The elasticity of elastomer display can use bullet Property modulus characterize.The modulus of elasticity of the material that some embodiments of the present invention use be about 1pa to about 1tpa or about 10pa extremely To about 1gpa or about 50pa to about 10mpa or about 100pa is to about 1mpa for about 100gpa or about 20pa, although being answered according to concrete Needs also can adopt these extraneous moduluss of elasticity.In some cases, the modulus of elasticity of material can be about 100mpa (MPa) or less.In other embodiments, the modulus of elasticity of material be about 75mpa or less, about 50mpa or less, About 25mpa or less, about 10mpa or less, about 8mpa or less, about 5mpa or less or about 2mpa or less.

Embodiment of the present invention provides microfluidic device, and it comprises assembly such as passage, valve and room, and described assembly is extremely Small part is comprised, embeds by the fast one or more layers of springform or level, or is formed or formed wherein by it.One Plant exemplary microfluidic body device and there is the reagent circulation road being formed in elastomer ground floor or reagent line.Described reagent circulation road Comprise to accommodate valve and room conduit.Microfluidic device also can have and is formed in the elastomer second layer adjoining with described ground floor Control passage or accommodate line.Additionally, microfluidic device can comprise to be formed at the elastomer threeth adjoining with the described second layer Sample flow channel in layer or sample wire.Described sample flow channel can comprise to accommodate valve and room conduit.Described control passage can be with Reagent circulation road accommodates valve and sample flow channel accommodates valve and is operatively connected.Described microfluidic device can comprise and reagent line fluid The reagent chamber of connection, and the sample room being in fluid communication with sample wire.Described reagent chamber and sample room can be by being formed at described bullet Reaction circulation road in gonosome third layer or response line and fluid communication with each other.Described response line can comprise interface valve.Described micro- Fluid means also can comprise the interface channel being formed in the 4th layer of the elastomer adjoining with described third layer.Described interface leads to Road can be connected with described reaction stream Entry Interface valve operability.

Embodiment of the present invention is also contemplated by manufacturing and the method using microfluidic device disclosed herein.For example, miniflow The operation of body device may include the one or more separate valves of unlatching, closes one or more interface valves and so that material is flow through solely Vertical valve simultaneously flows into one or more rooms, optionally carries out under stress.Technology may also include change seal line in pressure with Close separate valves, to close separate chambers, and change the pressure of parting line to open interface valve.First material of the first Room can flow Cross the interface valve opened and enter second Room, the wherein first material and the second material mixing contained therein or reaction.

It should be understood that example as herein described and embodiment are intended merely to example purpose, those skilled in the art can be according to it Suggestion is multiple to be improved or changes, and these are included in spirit herein and authority and scope of the following claims Interior.

Herein below corresponds to the original claims in parent application, is now incorporated to this as part of the specification Place:

1st, a kind of microfluidic device, it comprises

Pressure source；

Control line, it is with described pressure fluid communication；

Multiple valves, it is operated by described control line；With

Separate valves, it is located at described control line nearby and is between described pressure source and the plurality of valve.

2nd, the microfluidic device of item 1, wherein said separate valves build and are arranged in blocks fluid by described control line Flowing.

3rd, the microfluidic device of item 1, wherein said separate valves and second pressure fluid communication.

4th, the microfluidic device of item 1, wherein said control line includes the lock-in control line comprising check valve.

5th, the microfluidic device of item 4, wherein said check valve is arranged to stop fluid from described valve to described pressure source Flowing.

6th, the microfluidic device of item 5, it also comprises multiple rooms valve.

7th, the microfluidic device of item 6, wherein said control line provides to the plurality of valve and the plurality of room valve and starts pressure Power.

8th, the microfluidic device of item 1, wherein said separate valves can be used to stop the plurality of valve from being closed.

9th, the microfluidic device of item 1, it also comprises more than second valve by the second control line operation.

10th, the microfluidic device of item 9, wherein said second control line includes the second lock-in control comprising the second check valve Line.

11st, the method that operation has the microfluidic device of valve and control line, described control line has valve associated there Group, methods described includes:

Close described valve；With

Pressure is applied to described control line；The valve wherein cut out makes the described valve group being associated with described control line can not Operation.

12nd, the method for item 11, wherein closes described valve and includes:

Second pressure is applied to the second control line；With

Respond to described second pressure and close described valve.

13rd, the method for item 11, the wherein first valve group responds to described first control line applying first pressure and closes.

14th, the method for item 13, wherein said first valve group is blocked,

15th, the method for item 11, it also includes:

Keep the described second pressure applying to described second control line；With

Second pressure less than described first pressure is applied to described first control line, wherein with described second control line phase The valve group of association responds to the applying of described second pressure and closes.

16th, the method for item 15, wherein said valve group is blocked.

17th, a kind of microfluidic device, it comprises:

First valve；

Second valve；

Control line, it is in fluid communication with described first valve and described second valve；

Acute build up of pressure device, it is in fluid communication with described control line；With

Check valve, it is located near described control line and is located between described acute build up of pressure device and described second valve.

18th, the microfluidic device of item 17, the described control line between wherein said acute build up of pressure device and described first valve is no Check valve.

19th, the microfluidic device of item 17, wherein said first valve is arranged to be arranged in described microfluidic device Reative cell separates.

20th, the microfluidic device of item 17, wherein said second valve is arranged to first fluid input line and second fluid Input line separates.

21st, the microfluidic device of item 17, it also comprises the 3rd valve being in fluid communication with described control line.

22nd, the microfluidic device of item 21, wherein said 3rd valve can be used to stop fluid by mutually even with reative cell The flowing of fluid input line even.

23rd, the microfluidic device of item 21, it also comprises to build and be arranged to block fluid through dividing that described control line flows Every valve, wherein said separator valve is placed between described acute build up of pressure device and described 3rd valve along described control line.

24th, the microfluidic device of item 23, wherein said separator valve be set to accept from the second control line start pressure, Described second control line is in fluid communication with second pressure accumulator.

25th, the microfluidic device of item 24, wherein said second control line is independent of described control line.

26th, the microfluidic device of item 23, it is described that wherein said separator valve can be used to stop described 3rd valve to respond to Starting of control line and close.

27th, a kind of microfluidic device, it comprises:

Multiple reative cells, it is arranged as array layout, and each of the plurality of reative cell has and the plurality of reative cell One of the first valve of being in fluid communication and with one of the plurality of reative cell described the second valve being in fluid communication；

First control line, it is operable to start described first valve and described second valve；

Input line group, it is in fluid communication with the plurality of reative cell；

Multiple sample inlets, it is in fluid communication with described input line group；With

Check valve, it is arranged in described first control line.

28th, the microfluidic device of item 27, it also comprises to be in fluid communication with first pressure accumulator and be operable to hinder Fill in the first valve group of a part of described input line group.

29th, the microfluidic device of item 27, wherein said first valve group is in the downstream of described check valve.

30th, the microfluidic device of item 27, it also comprises:

Second input line group, it is in fluid communication with the plurality of reative cell；With

More than second sample inlet, it is in fluid communication with described second input line group.

31st, the microfluidic device of item 30, it also comprises can be by the second valve group of the second control line operation.

32nd, the microfluidic device of item 31, wherein said second control line is in fluid communication with second pressure accumulator, and can It is operated to block a part of described second input line group.

33rd, the microfluidic device of item 31, it also comprises the second check valve being arranged in described second control line.

34th, the microfluidic device of item 31, it also comprises to be arranged in described second control line and be operable to block Fluid is through the separator valve of described first control line flowing.

35th, a kind of microfluidic device, it comprises:

Multiple reative cells；

Multiple first input ports, each of wherein said multiple first input ports passes through one of more than first input line and institute State one or more of multiple reative cells to be in fluid communication；

Multiple second input ports, each of wherein said multiple second input ports passes through one of more than second input line and institute State one or more of multiple reative cells to be in fluid communication；

First pressure accumulator, it is in fluid communication with the first control line, and wherein said first control line is set off institute State more than first input line；

Second pressure accumulator, it is in fluid communication with the second control line, and wherein said second control line is set off institute State more than second input line；

First check valve, it is arranged in the institute between described first pressure accumulator and described more than first input line State in the first control line；With

Second check valve, it is arranged in the institute between described second pressure accumulator and described more than second input line State in the second control line.

36th, the microfluidic device of item 35, it also comprises to be operable to separate the valve of each of the plurality of reative cell Group.

37th, the microfluidic device of item 35, it also comprises to be operable to block described second control line and be arranged in Valve between described second pressure accumulator and described second control line.

38th, the microfluidic device of item 37, wherein said first control line can be used to start described valve.

39th, the microfluidic device of item 35, its subgroup also comprising to can be used to block described more than second input line Valve group.

40th, the microfluidic device of item 35, it also comprises the valve group that can be used to block manifold, and described manifold provides institute State the fluid communication between more than second input line.

41st, the method that operation has the microfluidic device of multiple valves and check valve, methods described includes:

Apply first fluid pressure to the control line of described microfluidic device；

Respond to the applying of described first pressure and close the plurality of valve；

Respond to the applying of described first pressure and close described check valve；With

Apply second fluid pressure to the second control line of described microfluidic device.

42nd, the method for item 41, it also comprises to reduce the described first fluid pressure being applied in described control line.

43rd, the method for item 42 is wherein after reducing the described first fluid pressure being applied in described control line, described many Individual valve remains off.

44th, the method for item 42, wherein reduces described first fluid pressure and includes not applying Fluid pressure to described control line.

45th, the method for item 41, wherein applies described second fluid pressure to described second control line and includes closing separator valve.

46th, the method for item 45, wherein said separator valve can be used to stop between the plurality of valve and described control line Fluid communication.

47th, the method for item 41, wherein said microfluidic device have multiple reative cells and with the plurality of reative cell in Each associated valve group.

48th, the method that operation has the microfluidic device of multiple input ports, methods described includes:

To one of the plurality of input port offer input fluid；

Start valve group to close the Part I of the input line being connected with the subgroup of the plurality of input port, wherein said Asia Group does not comprise the one of described of the plurality of input port；

Described input fluid is made to flow through the input line being connected with one of described in the plurality of input port；

Described input fluid is made to flow to the Part II of described input line through described input line；With

Close the second valve group to separate multiple reative cells.

49th, the method for item 48, wherein closes described second valve group and includes applying Fluid pressure to first pressure source.

50th, the method for item 49, wherein closes described second valve group and includes closing the discrimination providing fluid communication between input line Pipe.

51st, the method for item 49, wherein starts described valve group and includes applying Fluid pressure to second pressure source.

52nd, the method for item 48, wherein makes the described Part II that described input fluid flow to described input line include making institute State input fluid and flow through the manifold that fluid communication between described input line is provided.

53rd, the method for item 48, wherein started described valve group before closing described second valve group.

54th, the method for item 48, it also includes making described input fluid flow through the second input line group, described second input line group It is branched off into the described Part II of each of described input line.

55th, the method operating programmable microfluidic device, described microfluidic device has and the first input line group and second The reaction site array that input line group is in fluid communication, methods described includes:

Start the first valve group being operable to block described first input line group；

Start the second valve group being operable to the Part I of subgroup blocking the second input line group；

Sample is loaded into by described reaction site by the Part II of described second input line group；With

Start the 3rd valve group being operable to separate described reaction site.

56th, the method for item 55, wherein starts described first valve group and includes the first valve group described in locking.

57th, the method for item 55, is wherein started described first valve group and includes to first being in fluid communication with described first valve group Pressure source applies first pressure.

58th, the method for item 55, wherein starts described first valve group and starts described second valve group and carry out simultaneously.

59th, the method for item 55, is wherein started described 3rd valve group and includes to second being in fluid communication with described 3rd valve group Pressure source applies second pressure.

60th, the method for item 59, the starting after multiple samples are loaded into described reaction site of wherein said 3rd valve group Carry out.

61st, the method for item 55, wherein said sample through connection described second input line group Part II manifold from The Part I of described input line flow to the Part II of described second input line group, and the Part I of described input line does not comprise In the subgroup of described second input line group.

62nd, the method for item 55, is wherein not included in the input line in the described subgroup of described second input line group and setting Input port for receiving described sample is in fluid communication.

63rd, a kind of microfluidic device, it comprises:

The input port of predetermined quantity, each described input port is operable to receive one of multiple input fluids；

Multiple input fluid lines, one of each input port with described predetermined quantity of the plurality of input fluid line fluid Connection；

Valve group, each of described valve group is operable to close one of the plurality of input fluid line, wherein said valve The quantity of group is less than described predetermined quantity；

Manifold, its each fluid communication with the plurality of input fluid line；With

Second valve group, each of described second valve group is operable to close a part for described manifold.

64th, the microfluidic device of item 63, wherein said predetermined quantity is 12.

65th, the microfluidic device of item 63, it also comprises the first pressure source being in fluid communication with described valve group.

66th, the microfluidic device of item 65, it also comprises the second pressure source being in fluid communication with described valve group.

67th, the microfluidic device of item 63, the described part of wherein said manifold comprises fluid line, and described fluid line will be described Another of multiple input one of fluid lines and the plurality of input fluid line is connected.

68th, the microfluidic device of item 63, the quantity of wherein said valve group is less than the quantity of described predetermined quantity.

69th, the microfluidic device of item 63, each of wherein said multiple input fluid lines is branched off into multiple input fluid lines.

70th, the method operating programmable microfluidic device, described microfluidic device has reaction site array, and it is with first Input line group, the second input line group connect with the manifold fluid connecting described second input line group, and methods described includes:

Start the first valve group being operable to close described first input line group；

Start the second valve group being operable to the Part I of subgroup closing the second input line group；

Start the 3rd valve group being operable to make described manifold disable；

Described second valve group is started in revocation；

By the Part II of described second input line group, multiple samples are loaded in described reaction site；With

Start the 4th valve group being operable to separate described reaction site.

71st, the method for item 70, wherein starts described first valve group and includes the first valve group described in locking.

72nd, the method for item 70, wherein starts described first valve group and starts described second valve group and include to first pressure source Apply first pressure, described first pressure source is in fluid communication with described first valve group and described second valve group.

73rd, the method for item 72, wherein starts described 3rd valve group and includes second pressure source is applied second pressure, and described the Two pressure source is in fluid communication with described second valve group.

74th, the method for item 73, wherein said second pressure is less than described first pressure.

75th, the method for item 70, wherein revocation are started described second valve group and are included reducing and be applied to described first pressure source Described first pressure.

76th, the method for item 70, wherein starts described 3rd valve group and includes the 3rd valve group described in locking.

77th, the method for item 70, it also includes starting valve to stop the closing of the 5th valve group, and described 5th valve group can be operated In order to close the described Part II of described second group of input line.

78th, the method for item 77, wherein said valve and first pressure fluid communication, described first pressure source and described first Valve group is in fluid communication.

79th, the method for item 70, the Part I of each of wherein said second input line group is in fluid communication with input port, Described input port is set to receive one of the plurality of sample.

80th, the method for item 70, wherein said reaction site comprises multiple reative cells, and each of the plurality of reative cell is passed through Through hole is in fluid communication with the Part II of one of described second input fluid line group.

81st, the method for item 70, the Part I of each of wherein said second input line group and different input ports fluid Connection.

82nd, the method operating programmable microfluidic device, described programmable microfluidic device has and the first input line group The reaction site array being in fluid communication with the second input line group, methods described includes:

Start the second valve group being operable to separate described reaction site；

Described second valve group is started in revocation；

Multiple samples are loaded into described reaction site by the second input line group；With

Start described second valve group.

83rd, the method for item 82, wherein starts described first valve group and includes the first valve group described in locking.

84th, the method for item 82, wherein starts described first valve group and includes first pressure source is applied first pressure, and described the One pressure source is in fluid communication with described first valve group.

85th, the method for item 82, it is additionally included in while starting described first valve group and starts the 3rd valve group.

86th, the method for item 85, wherein starts described 3rd valve group and includes the 3rd valve group described in locking.

87th, the method for item 82, wherein revocation are started described first valve group and are included reducing and be applied to described first pressure source Described first pressure.

88th, the method for item 82, each of wherein said second input line group is in fluid communication from different input ports.

89th, the method for item 82, wherein starts described first valve group and starts described second valve group and carry out simultaneously.

90th, the method for item 82, wherein said reaction site is included in the elastomer layer be limited to described microfluidic device Reative cell.

91st, a kind of microfluidic device, it comprises:

Multiple reaction site；

First input line group, it provides the fluid between the first input port of predetermined quantity and the plurality of reaction site to connect Logical, wherein said first group of quantity is described predetermined quantity；

Second input line group, it provides the fluid between the second input port of predetermined quantity and the plurality of reative cell to connect Logical, each of wherein said second input line group comprises trunk portion and component, and described second group of quantity is few In described predetermined quantity；With

Programmable input device, it is operable to fill out using described first input line group or described second input line group Fill described reative cell.

92nd, the microfluidic device of item 91, wherein said multiple reaction site are arranged as array layout.

93rd, the microfluidic device of item 91, each of wherein said multiple reaction site comprise reative cell, described reative cell By through hole and one of described first input fluid line group fluid communication.

94th, the microfluidic device of item 91, each of wherein said multiple reaction site comprise reative cell, described reative cell By through hole and one of the component of described second input fluid line group fluid communication.

95th, the microfluidic device of item 91, the described component of each of wherein said input line group is from described trunk Part forms four fluid passages.

96th, the microfluidic device of item 91, wherein said programmable input device comprises to be operable to close described The valve group of one input fluid line group.

97th, the microfluidic device of item 96, wherein said valve group is in fluid communication with check valve, and described check valve can be used by operation With by described valve group locking in off position.

98th, the microfluidic device of item 91, wherein said programmable input device comprises valve group, and described valve group can be used by operation To close the described component of described second input fluid line group.

99th, the microfluidic device of item 98, wherein said valve group is in fluid communication with check valve, and described check valve can be used by operation With by described valve group locking in off position.

100th, the microfluidic device of item 98, it also comprises to be arranged in described valve group nearby and be operable to stop institute State the separate valves of valve group closing.

101st, the method that setting has the microfluidic device of multiple control lines, methods described includes:

Start the first control line；

Valve is placed in first state；

Afterwards, start the second control line being operable to that valve group is placed in the second state, be wherein in described first The valve of state stops described valve group from being placed in described second state.

102nd, the method for item 101, wherein said first state is to close.

103rd, the method for item 101, wherein said second state is to close.

104th, the method for item 101, wherein starts described first control line and includes applying first pressure to first pressure source.

105th, the method for item 104, wherein starts described second control line and includes applying second pressure to second pressure source.

106th, the method for item 105, wherein said second pressure source is independent of described first pressure source.

107th, a kind of method that setting has the microfluidic device of multiple control lines, methods described includes:

Then start the first state that the second control line sets up described microfluidic device by starting the first control line；With

By start described second control line then start described first control line set up described for the of fluid means Two-state.

108th, the method for item 107, wherein said first state include the first input line group being closed and with institute State the second input line group that the multiple reative cells arranged on microfluidic device are in fluid communication.

109th, the method for item 108, wherein said second state include with described microfluidic device on arranged described many Described first input line group and the described second input line group being closed that individual reative cell is in fluid communication.

110th, the method for item 108, wherein said second state also includes the valve group being closed, and it stops fluid stream Cross the manifold connecting described second input line group.

111st, the method for item 107, wherein said first control line and first pressure fluid communication.

112nd, the method for item 111, wherein said second control line and second pressure fluid communication.

113rd, the method for item 107, wherein said microfluidic device comprises elastomeric material.

114th, a kind of microfluid system, it comprises:

Carrier, it comprises:

Multiple first input ports；

Multiple first input lines, each of the plurality of first input line and one of the plurality of first input port fluid Connection；

Multiple second input ports；

Multiple second input lines, each of the plurality of second input line and one of the plurality of second input port fluid Connection；

First pressure source；With

Second pressure source

Dress microfluidic device on the carrier, described microfluidic device comprises:

Multiple 3rd input lines, each of the plurality of 3rd input line and one of the plurality of first input line fluid Connection；

Multiple 4th input lines, each of the plurality of 4th input line and one of the plurality of second input line fluid Connection；

The first control line with described first pressure fluid communication；

Check valve, it is operable to block at least a portion of described first control line；With

The second control line with described second pressure fluid communication.

115th, the microfluid system of item 114, wherein said multiple first input ports are operable to receive UP Source.

116th, the microfluid system of item 114, wherein said multiple second input ports are operable to receive UP Source.

117th, the microfluid system of item 114, it also comprises to be operable to block described second control line at least The second check valve dividing.

118th, the microfluid system of item 114, wherein said microfluidic device also comprises manifold, and it is by the institute in described carrier State multiple second input lines to be connected with the 4th input lines multiple described in described microfluidic device.

119th, the microfluid system of item 118, it also comprises the valve group being operable to the part blocking described manifold.

120th, the microfluid system of item 114, wherein said microfluidic device also comprises multiple reative cell groups, each reative cell One of each reative cell of group and the plurality of 3rd input line and one of the plurality of 4th input line are in fluid communication.

Claims

1. the method that setting has the microfluidic device of multiple control lines, methods described includes:

Start the first control line；

Valve is placed in first state；

Afterwards, start the second control line being operable to that valve group is placed in the second state, be wherein in described first state Valve stop described valve group be placed in described second state.

2. the method for claim 1 wherein that described first state is to close.

3. the method for claim 1 wherein that described second state is to close.

4. the method for claim 1 wherein that starting described first control line includes applying first pressure to first pressure source.

5. the method for claim 4, wherein starts described second control line and includes applying second pressure to second pressure source.

6. the method for claim 5, wherein said second pressure source is independent of described first pressure source.

7. a kind of method that setting has the microfluidic device of multiple control lines, methods described includes:

Then start the second shape that described first control line sets up described microfluidic device by starting described second control line State.

8. the method for claim 7, wherein said first state include the first input line group being closed and with described The second input line group that the multiple reative cells arranged on microfluidic device are in fluid communication.

9. the method for claim 8, wherein said second state include with described microfluidic device on arranged the plurality of Described first input line group and the described second input line group being closed that reative cell is in fluid communication.

10. the method for claim 8, wherein said second state also includes the valve group being closed, and it stops fluid stream Cross the manifold connecting described second input line group.

The method of 11. claim 7, wherein said first control line and first pressure fluid communication.

The method of 12. claim 11, wherein said second control line and second pressure fluid communication.

The method of 13. claim 7, wherein said microfluidic device comprises elastomeric material.