Summary of the invention
A purpose of the present invention is that providing a kind of biological detection system, being suitable for detection includes several target organism particlesLiquid sample.The biological detection system includes a capture device, passes through target organism particle described in the capture deviceIt can effectively be captured and can be separated easily from the capture device, is further processed for subsequent.
According to the present invention, a kind of biological detection system suitable for detecting the liquid sample comprising several target organism particlesIt is provided.The biological detection system include a capture device, and the capture device include a shell structure, one it is defeatedEntrance, a delivery outlet, a single-chip and one layer of active layer.
The shell structure includes a lower case and a upper housing.
The lower case has a bottom major surface and a lower binding face.The lower binding face is in a transverse directionOn in contrast to the bottom major surface and there is a lower groove, the lower groove extends downwardly and terminates at an inner bottom surface and packetIt includes one and is configured to sub- groove under the upper sub- groove for allowing the liquid sample to pass through and one.
The upper housing has a top main surface and a upper binding face.The upper binding face is in the transverse directionOn in contrast to the top main surface and there is a upper groove, the upper groove extends towards the top main surface and terminates at oneIt inner top surface and is configured to that the liquid sample is allowed to pass through.
The input port is arranged at the upstream of the lower groove and is used to for the liquid sample being introduced to described recessedIn the upper sub- groove of slot.
The delivery outlet is arranged at the downstream of the lower groove and on a longitudinal direction in contrast to the input port.
The single-chip is configured as matingly being embedded in the lower sub- groove, and includes a substrate and several dispersionsThe works of nano-scale, the works of the nano-scale of the dispersion is separately arranged in the longitudinal direction and eachIt is to be upwardly extended from the substrate and terminate at a top.
The active layer is formed in the top of each in the works of the nano-scale of the dispersion and is used to catchCatch the target organism particle.
Biological detection system of the invention, the works of the nano-scale of the dispersion are included the following steps by oneMethod is formed,
Step (a) provides one layer of etchable layer on the substrate；
Step (b), covers several regions on the etchable layer, and each of described region corresponds to the dispersionEach of the works of nano-scale the top；And
Step (c) etches downwards several non-obstructing regions of the etchable layer towards the substrate, in order to described in formationThe works of the nano-scale of dispersion,
Wherein, the step (b) includes that sub-step is as follows,
Step (b1) applies one layer of layer comprising photoresist on the etchable layer；
Step (b2) enables the layer comprising photoresist be exposed via a patterned shielding, and developsWith formed on the etchable layer one layer include the photoresist patterned layer, in order to define the non-obstructing region,And the photoresist is not provided in the non-obstructing area；
Step (b3) forms one layer of layer comprising silver catalyst on the non-obstructing region；And
Step (b4) lifts off the patterned layer comprising the photoresist, in order in the described non-of the etchable layerThe silver catalyst is left in shaded areas；And the step (c) includes that sub-step is as follows,
Step (c1) the obtained etchable layer will be soaked in comprising HF and H in the sub-step (b4)2O2Etching aqueous solution in, to enable the non-obstructing region of the etchable layer carry out silver-colored catalytic chemistry etching reaction, the silverCatalytic chemistry etching reaction can etch downwards towards the substrate, in order to the works to form the nano-scale of the dispersion, instituteThe works for stating the nano-scale of dispersion have because parallel lateral etch and caused by the mechanical strength through weakening；And
Step (c2) removes the silver catalyst from the etchable layer.
Biological detection system of the invention, the etchable layer are to be formed in one piece with the substrate.
Biological detection system of the invention, the works of the nano-scale of the dispersion are arranged in array.
Biological detection system of the invention, each of works of nano-scale of the dispersion is with one in instituteThe outer surface extended between substrate and the top is stated, and the outer surface is formed with number because of the parallel lateral etchA hole.
Biological detection system of the invention, the size that there is each of described hole range to fall in 500pm to 50nm.
Biological detection system of the invention, the single-chip are to be formed in one piece with the lower case.
Biological detection system of the invention, the input port include the lower input oral area for being formed in the lower case,And one be formed in the upper housing and be configured as and the lower upper input oral area that matches of input oral area, the outputIt mouthful include that a lower output oral area for being formed in the lower case and one are formed in the upper housing and are configured as and instituteState the upper output oral area that lower output oral area matches.
Biological detection system of the invention, the upper housing also have several conducting elements being set on the inner top surface,And the conducting element is separately arranged in the longitudinal direction and the fluid to deflect the liquid sample flows to.
Biological detection system of the invention also includes a separator, described to isolate from the single-chipThe works of the nano-scale of dispersion.
Biological detection system of the invention also includes a signal measurement module, comprising:
One signal measuring equipment；And
One the first metallic conductor and second metallic conductor, are set on the lower binding face and in the longitudinal direction sideIt upwardly extends and is spaced apart by the lower groove, it is each in first metallic conductor and second metallic conductorThere are two person's tools on the contrary and is electrically connected to the end of the signal measuring equipment.
Specific embodiment
The following describes the present invention in detail with reference to the accompanying drawings and embodiments.It, should before the present invention is described in detailPay attention in the following description content, similar element is indicated with being identically numbered.
Refering to fig. 1, a first embodiment of Fig. 2, Fig. 3 and Fig. 4, biological detection system according to the present invention are suitable for inspectionThe liquid sample comprising several target organism particles 6 is surveyed, and includes the separation of the electrode unit 2, one of capture device 1, oneDevice 5 and a signal measurement module 3.
The liquid sample, blood, lymph, urine, saliva etc. can be from an animal individuals or a human individualIt is obtained.The target organism particle 6 can be, such as cell, microorganism, protein, such as circulating tumor cellCore red blood cell (the fetal nucleated red blood of (circulating tumor cells, CTCs), fetusCells, fNRBCs), fetal trophoblasts (trophoblast) cell, virus, bacterium, antigen etc..In addition, the target organismParticle 6 can be obtained from plant, such as the tissue extractor object of plant.
The capture device 1 is used to capture the target organism particle 6, and including one shell structure 11, oneThe 16, single-chips 12 of delivery outlet of input port 15, one and one layer of active layer 123.
The shell structure 11 includes a lower case 111 and a upper housing 112.The lower case 111 and the upper housing112 engaged with one another define a runner 10.
The lower case 111 has a bottom major surface 1111 and a lower binding face 1112.The lower binding face 1112 existsIn contrast to the bottom major surface 1111 in one transverse direction, and there is a lower groove 101.The lower groove 101 extend downwardly andIt terminates at an inner bottom surface 1113 and is configured to the upper sub- groove 1011 and one for allowing the liquid sample to pass through including oneSub- groove 1012 under a.
The upper housing 112 has a top main surface 1121 and a upper binding face 1122.Binding face 1122 exists on thisIn contrast to the top main surface 1121 and with a upper groove 102 in the transverse direction.The upper groove 102 is towards the top main surface1121 extend and terminate at an inner top surface 1123 and be configured to that the liquid sample is allowed to pass through.The upper groove 102 and shouldLower groove 101 is engaged with one another and constitutes the runner 10.
The input port 15 is arranged at the upstream of the lower groove 101, and for the liquid sample to be introduced to the lower grooveOn this of 101 in sub- groove 1011.The input port 15 includes that oral area 151 and a upper input oral area 152 are inputted under one.It shouldLower input oral area 151 is formed in the lower case 111.Oral area 152 is inputted on this to be formed in the upper housing 112 and be configured as and be somebody's turn to doLower input oral area 151 matches.
The delivery outlet 16 is arranged at the downstream of the lower groove 101 and on a longitudinal direction in contrast to the input port15.The delivery outlet 16 includes that a lower output oral area 161 for being formed in the lower case 111 and one are formed in the upper housingIt 112 and is configured as and the lower upper output oral area 162 that matches of output oral area 161.
The upper housing 112 also has several conducting elements 131 for being set to the inner top surface 1123, and the conducting element 131 existsSeparately it is arranged on the longitudinal direction and the fluid to deflect the liquid sample flows to.The conducting element 131 is in herring-bone formFigure (herringbone pattern) is arranged in order to so that the liquid sample generates flow-disturbing effect in the runner 10(turbulent effect)。
The lower case 111 be by can be identical or different with material that the upper housing 112 is made material made by.Under thisIt is by semiconductor material, biological compatibility material or their combination in shell 111 and the upper housing 112 eachIt is independently made, will may be further described below.
Referring specifically to Fig. 2 and Fig. 3, which is configured as matingly being embedded in sub- groove 1012 under this, andThe works 121 of nano-scale including a substrate 124 and several dispersions is [for example, " nanostructure in nanometer " (" nano-on-nano structures")].The works 121 of the nano-scale of the dispersion on the longitudinal direction separately setting andEach is to upwardly extend from the substrate 124 and terminate at a top 130.
The works 121 of the nano-scale of the dispersion is arranged in array.The works of the nano-scale of the dispersionEach of 121 are formed such as a column type.In this embodiment, the single-chip 12 and the lower case 111 be one atFormed to shape.
The works 121 of the nano-scale of the dispersion is formed by the method included the following steps:
Step (a) provides one layer of etchable layer on the substrate 124；
Step (b), covers several regions on the etchable layer, and each of described region corresponds to the dispersionThe top 130 of each of the works 121 of nano-scale；And
Step (c) etches downwards several non-obstructing regions of the etchable layer towards the substrate 124, in order to described in formationThe works 121 of the nano-scale of dispersion.
Particularly, step (b) includes that sub-step is as follows:
Step (b1) applies one layer of layer comprising photoresist on the etchable layer；
Step (b2) enables the layer comprising photoresist be exposed via a patterned shielding, and developed [It is exactly, via light lithography imaging technique (photolithographic technique)], to form one on the etchable layerLayer includes the patterned layer of the photoresist, in order to define the non-obstructing region, and is not provided on the non-obstructing regionPhotoresist；
Step (b3) forms one layer of layer comprising silver catalyst on the non-obstructing region；And
Step (b4) lifts off the patterned layer that this includes photoresist, in order in the non-obstructing region of the etchable layerOn leave the silver catalyst.
Particularly, step (c) includes that sub-step is as follows:
Step (c1) the obtained etchable layer will be soaked in comprising HF and H in the sub-step (b4)2O2ErosionIt carves in aqueous solution, to enable the non-obstructing region of the etchable layer carry out silver-colored catalytic chemistry etching reaction.The silver catalytic chemistryEtching reaction can etch downwards towards the substrate 124, in order to the works 121 to form the nano-scale of the dispersion.Described pointThe works 121 of scattered nano-scale have because parallel lateral etch and caused by the mechanical strength through weakening；And
Step (c2) removes the silver catalyst from the etchable layer.
The etchable layer can be made by the material selected from following constituted group: semiconductor material, biology canBiocompatible material and their combination.One infinite example of the semiconductor material is silicon materials, such as silicon waferCircle.The biological compatibility material is such as, but not limited to dimethyl silicone polymer (Polydimethylsiloxane, PDMS), gathersMethyl methacrylate (Polymethyl methacrylate, PMMA), polycarbonate (polycarbonate, PC) or theyAny combination.In this embodiment, the etchable layer and the substrate 124 are to be formed in one piece.
Referring specifically to Fig. 3, each of works 121 of the nano-scale of the dispersion is with one in the substrateThe outer surface 120 extended between 124 and the top 130, and the outer surface 120 is formed with number because of the parallel lateral etchA hole 122, so that each of works 121 of the nano-scale of the dispersion is allowed to have the machinery through weakening strongDegree.Described hole 122 is also formed on the top 130 of each of works 121 of the nano-scale of the dispersion.It is describedThe size that there is each of hole 122 range to fall in 500pm to 50nm.The works 121 of the nano-scale of the dispersionEach of the length and width ratio with 2 μm of width below and 5 or more.The works 121 of the nano-scale of the dispersionEach of there is range to fall in 30 to 50% porosity and range falls in 200 to 800m2/cm3Specific surface area.Compared withGoodly, so high specific surface area makes have the charge inducing opposite with the charged charge of the target organism particle 6 canIt is generated and is gathered in the surface of each of works 121 of the nano-scale of the dispersion, so that the target organismParticle 6 more effectively can be attracted and be captured by the works 121 of the nano-scale of the dispersion.
In Fig. 8 and Fig. 9 it is shown be through this embodiment in method as described above be formed by the dispersionNano-scale works 121 an example.
The active layer 123 is formed in the top 130 of each in the works 121 of the nano-scale of the dispersion and is used forCapture the target organism particle 6.The active layer 123 is to be intended to capture by that can specifically bind (specific binding)The target organism particle 6 action material made by, and the action material can be antigen-like material, antibody material, victory peptideMaterial, protein material or their any combination.In order to strengthen the capture effect of the active layer 123, the institute to be capturedStating target organism particle 6 can be incorporated in biotinylated antibody (biotinylated antibody), and the active layer 123It is streptavidin (streptavidin) the material institute by having specific binding interaction between biotinIt is made.The active layer 123 is the silane via the works 121 of the nano-scale of the dispersion obtained in step (c)Change reaction (silylation), is then coated on the action material every in the works 121 of the nano-scale of the dispersionOne top 130 and be formed.
The described hole 122 on the top 130 in the works 121 of the nano-scale of the dispersion each can lead toIt crosses following method and is formed, for example, extending erosion time so that each in the works 121 of the nano-scale of the dispersionThe specific surface area on the top 130 of person can be enhanced, so increase be formed in the dispersion nano-scale works 121 inThe surface area of the active layer 123 on the top 130 of each is so that promote the combination effect of the active layer 123.Further, since thisSpecific surface area through improving, the induction assembled on the top 130 of each in the works 121 of the nano-scale of the dispersionCharge can also be increased, so that the effect for capturing the target organism particle 6 can be promoted further.
The electrode unit 2 includes the first electrode 21, Yi Jiyi for being set to the bottom major surface 1111 of the lower case 111The second electrode 22 of a top main surface 1121 for being set to the upper housing 112.In order to enhance institute included in the liquid sampleThe movement of target organism particle 6 towards the active layer 123 is stated, and prevents unwanted substance included in the liquid sample heavyProduct is on the active layer 123, during the liquid sample flows through runner 10, the first electrode 21 and the second electrode 22A voltage can be alternately imposed, to allow the capture device 1 between a trap mode and a non-trap modeIt is switched.Under the trap mode, a non-uniform electric field can be generated in the first electrode 21 and the second electrode 22 itBetween to allow dielectrophoresis phenomenon (dielectrophoresis) to occur, and make the target included in the liquid sampleBiomone 6 and the unwanted substance are mobile towards the active layer 123 by dielectrophoretic force attraction, and make the targetBiomone 6 is thus incorporated into the active layer 123, as shown in Figure 7.Under the non-trap mode, one in trap modeGenerated in the opposite non-uniform electric field of electric field can be generated between the first electrode 21 and the second electrode 22, causeThe dielectrophoretic force contrary with the dielectrophoretic force generated in trap mode generates, and makes included in the liquid sampleThe unwanted substance far from the active layer 123, and the target organism particle 6 is then persistently incorporated into the active layerOn 123.Be repeated several times switching circulation after, the target organism particle 6 can thus be effectively bonded to and concentrate on the workWith on layer 123.In a variation aspect of the present embodiment, which is not arranged in the shell structure 11,The first electrode 21 is applied with an electric current so that the lower case 111 and the single-chip 12 are electrically polarized(electrically polarized) and generate electric dipole moment (electric dipole moment), to allow the targetBiomone 6 is incorporated in on the active layer 123.
The signal measurement module 3 includes a signal measuring equipment 32, and is arranged on the lower binding face 1112One the first metallic conductor 31 and second metallic conductor 33.First metallic conductor 31 and second metallic conductor 33 are at thisExtend on longitudinal direction and is spaced apart by the lower groove 101.There are two opposite and electrical connections for first metallic conductor 31 toolTo the end of the signal measuring equipment 32 311.There are two second metallic conductor 33 tools on the contrary and is electrically connected to signal measurementThe end 331 of device 32.
The liquid sample that detection includes the target organism particle 6 is used in the biological detection system according to the present inventionBefore, an a pair of end 311 for measuring signal and being applied respectively to first metallic conductor 31 from the signal measuring equipment 32And the one end 331 of second metallic conductor 33, and a pair of of feedback signal respectively from first metallic conductor 31 anotherEnd 311 and another end 331 of second metallic conductor 33 are received by the signal measuring equipment 32.With receivedThe feedback signal as background signal data.
The biological detection system of the invention is then used to the liquid sample that detection includes the target organism particle 6,And receive another pair feedback signal by repeating program as described above, and using this to feedback signal as changing signal numberAccording to.Substantive signal data is then obtained by the way that the variation signal data is deducted the background signal data.The target is rawThe quantity and type of object particle 6 are via immunofluorescence dyeing technology (immunofluorescence stainingTechnique) arrange in pairs or groups fluorescence microscope and be analyzed.A series of liquid of target organism particles 6 comprising different numberSample is detected via program as described above, to establish a database.
Later, the liquid sample to be detected comprising the target organism particle 6 can via program as described above and byDetection, to obtain variation signal data.The variation signal data is brought to be compared with the data in the database, in order to trueThe quantity and type of the fixed target organism particle 6.
The signal measuring equipment 32 included by the embodiment is one and uses differential signal (differentialSignal) as the vector network analyzer (vector network analyzer, VNA) for measuring signal.
Referring specifically to Fig. 4, which is the nano-scale to isolate the dispersion from the single-chip 12Works 121.The separator 5 includes one for shaking the single-chip 12 with the structure for the nano-scale for destroying the dispersionThe oscillator 51 of object 121, in order to the works 121 to separate the nano-scale of the dispersion from the shell structure 11.In the implementationIn example, which is a sonicator.Since the works 121 of the nano-scale of the dispersion has because of shapeDescribed hole 122 on the outer surface Cheng Yuqi 120 and caused by the mechanical strength through weakening, so they can pass through the concussionDevice 51 and be destroyed easily.
Referring again to Fig. 1 and Fig. 4, include several target organisms when biological detection system according to the present invention is used to detectionWhen the liquid sample of particle 6, when flowing through the runner 10, the fluid flow direction of the liquid sample can be led the liquid sample by describedThe deflection of stream part 131 and cause the liquid sample to generate flow-disturbing effect in the runner 10, and increase the target organism particle 6 andIt is formed in the contact in the works 121 of the nano-scale of the dispersion between the active layer 123 on the top 130 of each.TogetherWhen, during the liquid sample flows through runner 10, the first electrode 21 of the electrode unit 2 and 22 meeting of second electrodeA voltage is imposed, alternately to allow the capture device 1 quilt between a trap mode and a non-trap modeSwitching, and the target organism particle 6 is effectively incorporated in and is concentrated on the active layer 123.It is trapped inThe target organism particle 6 on the active layer 123 can be measured by the signal measurement module 3.Finally, measuringAfterwards, capturing has the works 121 of the nano-scale of dispersion of the target organism particle 6 can be by using the separator 5And it is separated from the single-chip 12.
Refering to Fig. 5, a second embodiment of biological detection system according to the present invention is shown be analogous to this firstEmbodiment, in addition in this second embodiment, which defines several runners 10 and including several by accordinglyThe single-chip 12 being set in the runner 10.
Refering to Fig. 6, the first embodiment and second embodiment can further comprise one and be set on the separator 5The cutter 4 of trip.The cutter 4 is to isolate the portion of monocrystalline for capturing and having the target organism particle 6 from the shell structure 11Piece 12.One infinite example of the cutter 4 is a UV laser cutter.The part cut down by the cutter 4Single-chip 12 is then shaken by the oscillator 51, has the target organism in order to isolate capture from the single-chip 12The works 121 of the nano-scale of the dispersion of particle 6.
Only as described above, only the embodiment of the present invention is all when cannot be limited the scope of implementation of the present invention with thisIt is all still to belong to the present invention according to simple equivalent changes and modifications made by scope of the invention as claimed and description and coverIn the range of.