CN106918694A - Using the chip apparatus for joint inspection Diagnostic Value of Several Serum Tumor Markers of hydrophobic substrate - Google Patents

Abstract

The present invention relates to a kind of chip apparatus for joint inspection Diagnostic Value of Several Serum Tumor Markers using hydrophobic substrate, belong to analysis testing field.PDMS be dimethyl silicone polymer such as making the substrate that general type more than ten plants tumor markers joint-detection micro-fluidic chip, have a clear superiority, but there is also and some need the problems that overcome；This case is directed to the problem.This case main points are, substrate selectes the PDMS with ecosystem surface, and its neighbor positions fixes installing miniature ultrasonic transducer units in the test liquid stream terminal of the micro-fluidic chip, with ultrasonic wave reduction interfacial tension, while strong absorbabilities of the utilization PDMS to ultrasonic wave, reach ultrasonic intensity rapid decrement in short distance, so as to form interfacial tension difference at the two ends of the chip, difference offer is a kind of to drive the strength that test liquid stream flows along hydrophobic capillary channel to the terminal direction, the strength with Micropump its mechanicalness pumping strength for including in structure while intercouple, Collaboration.

Description

Using the chip apparatus for joint inspection Diagnostic Value of Several Serum Tumor Markers of hydrophobic substrate

Technical field

The present invention relates to a kind of chip apparatus for joint inspection Diagnostic Value of Several Serum Tumor Markers using hydrophobic substrate, belong to analysis test neck Domain.

Background technology

Tumor markers (tumor marker, TM) refers in the generation and breeding of tumour, to be produced in itself by tumour cell It is raw or by body to tumour cell react and produce, reflection tumour exist and growth a class material, including protein, Hormone, enzyme (isodynamic enzyme) and oncoprotein etc..Tumor markers in chemical examination blood samples of patients or body fluid, can be in cancer screening Early detection tumour, and observe the curative effect of oncotherapy and judge patient's prognosis.Clinically conventional tumor markers has at present： (1) alpha-fetoprotein (AFP) is the mark of the tumours such as primary carcinoma of liver, carcinoma of testis, oophoroma；(2) carcinomebryonic antigen (CEA) is digestion The mark of the tumours such as system tumor, lung cancer, breast cancer；(3) CA125 (CA125) is the mark of the tumours such as oophoroma； (4) CA153 (CA153) is the mark of the tumours such as breast cancer；(5) CA19-9 (CA19-9) is digestive system tumor Mark；(6) CA724 (CA724) is the mark of the tumours such as stomach cancer, oophoroma；(7) carbohydrate antigen 242 (CA242) It is the mark of digestive system tumor；(8) CA50 (CA50) is the mark of the tumours such as digestive system tumor, breast cancer, lung cancer Thing；(9) CYFRA21-1 (Cy211) is the mark of the tumours such as non-small cell lung cancer；(10) neuronspecific enolase (NSE) It is the mark of the tumours such as ED-SCLC, neuroendocrine tumor；(11) PSA (PSA) is prostate cancer Tumor markers；(12) human chorionic gonadotrophin (HCG) is swollen ECC, trophoblastic tumor (suede cancer, vesicular mole) etc. The mark of knurl；(13) thyroglobulin (TG) is the mark of thyroid cancer；(14) ferritin (SF) is digestive system tumor, liver The mark of the tumours such as cancer, breast cancer, lung cancer；(15) B2M (β 2-MG) is in chronic lymphocytic leukemia, lymph Raised in the patient body fluids such as knurl, myeloma, lung cancer, thyroid cancer, nasopharyngeal carcinoma；(16) squamous cell antigen (SCC) be cervical carcinoma, The tumor markerses such as lung squamous cancer, the cancer of the esophagus.The tumor markers overwhelming majority for clinically detecting at present is not only present in malignant tumour In, exist in benign tumour, embryonic tissue, even in normal structure.Therefore, tumor markers has dynamic chek and multinomial Joint inspection is more valuable.So for numerous tumor markerses, clinically how to selectDifferent tumours understands some phases To special tumor markers, such as CA153 often appears in breast cancer；CEA often appears in intestinal cancer, stomach cancer；CA19-9 is often appeared in Intestinal cancer, cancer of pancreas：CA125 often appears in oophoroma etc..Clinician can be according to the different mark of different tumor examinations. Same tumour or different types of tumour can have one or more tumor markers exceptions；Same tumor markers can be in difference Tumour in occur.To improve the additive diagnostic value of tumor markers and determining which kind of mark can be supervised as the follow-up after treatment Index is surveyed, tumor markers joint-detection, the complementary tumor markers group of several sensitivity of reasonable selection, specific performance can be carried out Into best of breed, joint-detection is carried out.In general the joint-detection of tumor markers can improve the accuracy to diagnosing tumor.

Only with regard to Diagnostic Value of Several Serum Tumor Markers its joint-detection background of related itself general picture or overview for, may refer to Lower Chinese invention patent application case：CN200410041175.3、CN200510026780.8、CN200610040051.2、 CN200910064647.X。

Only for the microfluidic chip technology overall general picture of itself, famous micro-fluidic expert Mr. Lin Ping Cheng is may refer to soon Before the monograph " diagram Microfluid based Lab on a chip " that goes out, the monograph publishes via Science Press, and the monograph is for micro-fluidic Past of technology, now, and, vision of the future etc. aspect, suffer from detail, be deep into long discussion of detail.

So, the focal issue that this case that to have a talk below pays special attention to.

The basic framework of micro-fluidic chip, including the substrate and the cover plate that fits together therewith of small fluid course are etched with, Small fluid course on the substrate, before upper cover plate is assembled, it is apparent on see to be exactly some micro-channels, be when thereon After covering cover plate, just real closure forms the small fluid course, and the conduit inner surface of the micro-channel is together with surround this The part cover plate of micro-channel constitutes described small fluid course together；So, it is clear that this after assembling is completed is small Fluid course, the major part of its inner surface area is the inner surface area of that micro-channel, in other words, in the micro-channel The state or property on surface substantially determine the integrality or property of the small fluid course；Therefore say, this is built in base The inner surface state or inner surface property of the micro-channel on piece are key factors；In principle, it is any to keep or protecting substantially The material of its solid forms is held, can be used to make substrate and cover plate, such as, the material that can act as substrate and cover plate can be with It is monocrystalline silicon piece, quartz plate, sheet glass, high polymer such as dimethyl silicone polymer, polymethyl methacrylate, makrolon etc. Deng；Certainly, the selection of substrate and the selection of cover plate can be with identical, it is also possible to differ；From material consumption, manufacture difficulty and Application popularization prospect etc. aspect from the point of view of, between these materials exist not small difference, the especially selection of that substrate, influence compared with Greatly.

In various substrate making materials, dimethyl silicone polymer, i.e. PDMS, comparatively very easily shaping, so Substrate on make that micro-channel is extremely simple, and the lower cost for material makes substrate with the polydimethyl siloxane material, Micro-channel is suppressed or etched thereon, and the cover plate made with glass or the cheap material such as polypropylene or other plastic sheets is engaged, It is seemingly a kind of more satisfactory selection；Certainly, patch material can also select to use cheap polydimethyl siloxane material： So, this substrate selection is the scheme of polydimethyl siloxane material, and material is extremely cheap, makes extremely simple, seems and should also be as It is extremely easy to popularize, promotes.

But, thing is really not so simple.

First, this polydimethyl siloxane material, that is, the material that the letter PDMS that abridges is referred to, itself are a kind of strong Hydrophobic material, micro-channel is built on this material, if not carrying out the modified operation for the micro-channel surface, then, After overall assembling is completed, that is, after covering cover plate, because of structure in the micro-channel its inner surface occupy most liquid circulation The inner surface in road, then, the PDMS micro-channels inner surface its strong hydrophobic property is deciding factor, and it can cause class Be similar to the aqueous solution the fine liquid stream of polar liquid by becoming very difficult, its flow resistance is big, or even general Micropump is all It is difficult to promote, certainly, if cover plate also selects to use the PDMS material, then, problem is substantially identical, similar； Therefore, among prior art, modification is modified particular for the micro-channel inner surface on the PDMS material, is necessary Operation；So, this is pretty troublesome for the modified operation of PDMS micro-channel inner surfacesThat falls nor this problem, structure It is another problem into serious technical puzzlement：PDMS polymer molecules tool inside this PDMS material substrate its body phase There is the characteristic for diffusing to the surface automatically, migrating, this substrate body phase inside PDMS polymer molecules are diffused to the surface, moved automatically The characteristic of shifting, by cause by that micro-channel of surface modifying and decorating its inner surface it is modified after state can not maintain foot Enough long time, being held time for micro-channel its inner surface state after that is surface-modified be substantially only sufficient in completion laboratory The time of portion's test experiments needs；In other words, PDMS micro-channel inner surfaces of or surface modification modified by surface, its After modified or say the surface state formed after modification can not be lasting, but soon automatically tend to or say become surface again and change Surface state before property, returns to the strong hydrophobic surface state of that script in the shorter time, then, just think, Such micro-fluidic chip can largely make, mass storage, be widely popularized, and answer is it is obvious that is, impossible. Micro-channel on this PDMS material, does not do if surface modification, and the fine liquid stream of polar solvent similar to the aqueous solution cannot pump Send and pass through, chip also cannot just be used；And if having done surface modification, its state after modifying cannot be persistently kept again, also Being equally cannot popularization and application.

So, how to accomplish that substrate can either be made using cheap PDMS material, and table in the micro-channel can be released Face decorating state cannot persistently, chip cannot largely make, largely lay in so that be widely popularized it is such a make this area it is numerous specially The puzzlement that industry personnel are entangled with for a long time, the highly difficult problem that exactly one its obvious technology barrier can not despise.

Be present many year in the highly difficult problem, so far, not yet properly settled.

Second, the PDMS material of non-surface modification, above it is stated that, its surface is strongly hydrophobic, this strong hydrophobic Material surface and also another problem, that is, this strong hydrophobic PDMS surfaces can adsorb large biological molecule, and And, the depression that these adsorbed large biological molecules can also be further further on PDMS surfaces gradually falls into gradually deep, directly It is trapped within the body phase of PDMS substrates to heavy, in fact, this process is partly also due to inside PDMS material body phase Polymer molecule have diffuse to the surface, caused by travel motion；Such case, it is also possible to explained from another angle, i.e. Continuously from inside PDMS body phases to its diffusion into the surface, those polymer molecules of migration, the result of its motion, be by Gradually those are involved within the body phase of PDMS substrates by the large biological molecule of adsorption, briefly, these are inhaled Attached large biological molecule is exactly to be swallowed up by PDMS substrate body phases；So, this PDMS substrates body phase swallows up large biological molecule Phenomenon, the influence caused by it necessarily causes to be related to the severe deviations of all kinds of test data of experiment of large biological molecule.

As described above, the problem of PDMS substrates is, its not only adsorption large biological molecule, and swallow up large biological molecule, So, as the large biological molecule of experiment test object, its disappearance will not stop because surface saturation is adsorbed, but, no It is disconnected adsorbed, also constantly swallowed up.

On PDMS substrates in related experiment test process its body phase constantly swallow up test associated biomolecule macromolecular phenomenon, separately It is to say that one kind is explained, there are substantial amounts of Minute pores in PDMS body phases, associated biomolecule macromolecular by after adsorption, depression Into these Minute pores, and then swallowed up；However, inventor thinks, those can allow the air point of miniature scale Son clamp-ons the Minute pores therebetween, and being not equal to them also can directly allow that the large biological molecule of relative large scale enters, and two Person's difference on yardstick is huge, must not make sweeping generalizations.Explanation is bypassed, in any case, the biology of object is analyzed as dependence test Macromolecular is adsorbed by PDMS substrate micro-channels inner surface, and then is constantly swallowed up by PDMS substrate body phases, and this is known objective The phenomenon of presence.

In order to prevent this PDMS substrate bodies relative to the effect of swallowing up of large biological molecule, can be from containment PDMS surfaces opposite The absorption of thing macromolecular is addressed, method be chemically modified aiming at the PDMS material surface it is modified, for For PDMS is for the situation of substrate material, exactly the surface of described micro-channel part is chemically modified it is modified, by changing The micro-channel inner surface of modification is learned, its absorption to large biological molecule can be contained, and then avoid large biological molecule quilt PDMS substrate body phases are swallowed up；But, or that old problem, that is, the chemical modification on PDMS material surface changes Surface state after property cannot persistently keep, the polymer molecule inside the PDMS substrate body phases its diffuse to the surface automatically, The process of migration, soon can become that again script strong and dredge by the micro-channel inner surface state that surface chemical modification is modified Water and the state of strong adsorption large biological molecule, in other words, no matter how professionals in the field turn from side to side, the PDMS Substrate its micro-channel inner surface is always rapidly to strong hydrophobic surface state evolution.

So, how can either obtain that PDMS material price is extremely cheap, substrate makes extremely easy benefit, can reach again Growth stage contains absorption process of the PDMS substrate micro-channel inner surfaces to large biological molecule, and then prevents PDMS substrate body phases The effect of swallowing up to large biological molecule so that related chip manufactured goods are able to maintain that a prolonged enough, rational shelf-life, It is exactly a very thorny problem.The problem makes the numerous specialties in this area as another problem addressed above, equally Personnel are entangled with, perplex for a long time, and the problem is equally the highly difficult problem that its obvious technology barrier can not despise.The hardly possible Also be present many year in topic, so far, also not yet properly settled.

Third, as described above, the PDMS micro-channel inner surfaces are strongly hydrophobic, and targetedly surface chemical modification or come to the surface Modification is learned to be difficult to again persistently, therefore, actually can only its surface is modified or surface modification after surface state still belong to effective short It is used within phase；If have passed through that ofer short duration period of validity, and still by force if use, due to surface shape State already again near hydrophobic state, drives test liquid stream then to certainly exist than larger flow resistance using usual Micropump, So, can only just force test liquid to flow to target direction by increasing Micropump pump power and pumping pressure to flow, as described above, This PDMS material is soft, and test liquid stream is pumped with too high, machinery pumping pressure, and it enters will to cause the substrate Sample end includes that the micro-channel of sample introduction end near zone occurs bubbling, expanded, distortion, deformation, also, this high-pressure situations Under, micro-channel and its periphery in the sample introduction end and its near zone are also susceptible to the stripping between substrate and cover plate, this feelings Under shape, sample solution will everywhere flow over into the crack of the appearance between the substrate of formation after the stripping and cover plate, this reality Border causes the damage of the micro-fluidic chip；Certainly, if the surface is modified or surface modification is not in place if originally, also result in There is said circumstances within the of short duration usual term of validity；It is above-mentioned in the case where liquid stream driving is carried out using additional Micropump merely The problem exists all the time.If as described above, do not did that any surface is modified completely or the pre action such as surface modification, So, the above-mentioned problem will be even more serious, even without occur sample introduction end and its micro-channel bubbling described near zone, it is expanded, Between distortion, deformation and substrate and cover plate the problems such as peel off, merely because the flow resistance is excessive, using high pressure Micropump also not Test liquid stream must be driven to be marched forward towards terminal side.

The content of the invention

The technical problems to be solved by the invention are to provide a package solution, while solving three for addressing above The a series of problem for actually mutually involving together of aspect, also, it is a kind of new that the solution is applied into structure Crowd's physical examination it is blanket can be directed to ten kinds than more typical primary tumor mark carry out simultaneously examination, while detection it is micro- Fluidic chip device.

The present invention solves the technical problem by following scheme, and the device that the program is provided is a kind of use using hydrophobic substrate In the chip apparatus of joint inspection Diagnostic Value of Several Serum Tumor Markers, program main points are that the structure of the device includes micro-fluidic chip, the miniflow Controlling the structure of chip includes installing bonded to each other substrate and cover plate together, and the substrate and cover plate are plate object or sheet The channel structure formed via mould pressing process or etching technics is contained in thing, that face towards the cover plate of the substrate, bonded to each other The substrate being installed together has been built into the micro-fluidic chip containing pipeline configuration, the locations of structures of the pipeline jointly with the cover plate Positioned at the substrate and cover plate juncture area bonded to each other, the two ends of the pipeline respectively with the sample introduction end of the micro-fluidic chip and Terminal is connected, and the sample introduction end is the injection end of the micro-fluidic chip sample solution, and the terminal is that the actual sample introduction of the micro-fluidic chip is surveyed The terminal that sample solution flows in its chip during examination, the terminal is located remotely from each other with the sample introduction end, between the terminal and the sample introduction end Distance between 3 centimetres and 10 centimetres, on diverse location in the pipeline in order or backward be equiped with working electrode and To electrode and reference electrode, the order refers to the reference electrode its locations of structures closer to the terminal location, described Backward refers to the reference electrode locations of structures closer to the sample introduction end position, the working electrode by conductive electrode and The gold size sensitive membrane for having embedded tumor markers antibody being attached on the conductive electrode is constituted, and the construction of the pipeline is presented in parallel Construction, the pipeline in parallel construction is made up of 15 lateral parallel connections, the pipeline that parallel construction is presented its Appearance profile is similar to the profile of parallel circuit, and the quantity of the working electrode is 15,15 installings of working electrode Position is located in 15 laterals respectively, and, in the sensitive membrane structure of its top layer gold size of 15 working electrodes Tumor markers antibody is respectively the 15 kinds of tumor markers antibody materials that can be specifically bound to tumor markers antigen, and this ten Five kinds of antibody materials be respectively tumor markers antibody A FP, CEA, CA242, CA125, CA199, CA153, CA724, CA50, NSE, CYFRA21-1, FPSA, TPSA, β-hCG, SCCA and β 2-MG, the antigen is the antigen of broad sense, described Antibody is the antibody of broad sense, and the working electrode its material is that argent material, gold material, carbon material or thermal decomposition are conductive Macromolecule material, the working electrode its pattern is presented column, sheet or thread, and the substrate its material is dimethyl silicone polymer Material, its surface of the substrate is the surface of primary form, the surface of the primary form its be intended to refer to not by any surface The surface of the primary form of the material of chemical modification or any surface chemical modification, the structure of the device also includes miniature ultrasonic Transducer, and, higher-order of oscillation electric signal transmission cable, one end and the miniature ultrasonic of the higher-order of oscillation electric signal transmission cable Wave transducer links together, the miniature ultrasonic transducer units attach be installed in the micro-fluidic chip cover plate or substrate it is neighbouring The position of the terminal；The miniature ultrasonic transducer units its major functions is when the actual sample introduction of micro-fluidic chip is tested, using it The ultrasonic wave launched reduces the interfacial tension between sample solution and the inwall of its inner passage of micro-fluidic chip, can It is enough compatible also, poor with the distance between the miniature ultrasonic transducer units installation position using the sample introduction end and the terminal Difference on different and its ultrasonic intensity for being experienced, its interfacial tension of sample introduction end described in induced synthesis and its boundary of the terminal Difference between the tension force of face, interfacial tension difference between the micro-fluidic chip two ends can the two ends of the micro-fluidic chip it Between form pressure gap, the pressure gap can drive sample solution to the end flow；The miniature ultrasonic transducer units its functions Also include the ultrasonic wave launched with it check contained large biological molecule in sample its in its inner passage of the micro-fluidic chip Absorption on inner surface, and then check swallow up work of its body phase of the substrate of the dimethyl silicone polymer material to the large biological molecule With；Softness simultaneously has its function of the substrate of the dimethyl silicone polymer material of elasticity including with its property to the strong absorption of ultrasonic wave Matter, is absorbed strongly to ultrasonic wave, and thereby in micro-fluidic chip terminal to the limited short distance between the sample introduction end Within realize the rapid decrement of ultrasonic intensity；And, Micropump, the Micropump is connected with the sample introduction end；The function of the Micropump is, Interfacial tension between the inwall of the sample solution with its inner passage of micro-fluidic chip is reduced by the ul-trasonic irradiation, phase Between under the increased precondition of compatibility, between the mechanicalness pumping strength of the Micropump is come and the ultrasonic wave is induced two ends Interfacial tension difference its driving force brought support mutually, adjust mutually, be mutually coupled, converged in the way of Collaboration Being polymerized to one drives test liquid to flow to the strength of the terminal direction flowing.

The tumor markers antibody is the antibody of broad sense, and the antibody of the broad sense is referred to possessing antibody function or is functionally similar to Can be combined with the various involved tumor markerses of corresponding clinic and forming immune complex or immune compound in antibody The material of the analog of thing；The tumor markers antigen is the antigen of broad sense, and the antigen of the broad sense is referred to can be using corresponding Antibody or be functionally similar to antibody material carry out enzyme mark detection it is various clinical involved the need for differentiate, the tumor-marker of detection Thing.

The gold size sensitive membrane is to be sufficiently mixed uniformly shitosan gold size solution and tumor markers specific antibody solution, is used Point sample instrument point sample coats specified structure position, and forms its drying and forming-film.Tumor markers in the gold size sensitive membrane Specific antibody is the tumor markers antibody of horseradish peroxidase or glucose oxidase mark, the gold size sensitive membrane It has been included as fixing above-mentioned each tumor markers specific antibody and introducing complementary medium therein, the complementary medium is for example Shitosan, cellulose acetate, gelatin be therein a kind of or their mixture.

The word of the Micropump one art-recognized meanings of itself are known for the professional in micro-fluidic chip field.

The Micropump both can be the Micropump of external form；The Micropump can also be done into or say inside the embedded micro-fluidic chip The Micropump of the built in version of its sample introduction end structure position or the sample introduction end Near-neighbor Structure position.

For example miniature piezoelectric pump of the Micropump, miniature peristaltic pump or miniature air driven pump.

The pipeline in the microfluidic chip structure includes the lateral, and its internal diameter size may each be arbitrarily selected Size, but, for prepare liquid sample and the consideration of the aspect such as reagent loss is reduced less as far as possible, described in the pipeline includes The passage of the preferred capillary level of lateral, the passage of the capillary level implies that the interior of the capillary on its internal diameter and ordinary meaning The suitable passage in footpath.The shape of cross section of its inner passage of capillary can be arbitrary shape, the shape of cross section example Such as circular, oval, square, rectangle, bar shaped, naturally it is also possible to be arbitrarily the presence of the linear of bending, also, the hair With the extension of pipeline, the shape of cross section of different parts can also allow to be different shapes the interior shape of tubule.Only with regard to hair For the word of tubule one, its art-recognized meanings is known.

What is be related in structure is the electrode of microsize to electrode and reference electrode, and its electrode shape may each be any choosing Fixed shape, the arbitrarily selected shape such as column, sheet, strip or thread etc..It is described to electrode and the ginseng Art-recognized meanings than the electrode vocabulary of itself are known.

It is public only for professional of the word of ultrasonic transducer one art-recognized meanings of itself for ultrasonic technology field Know.

Various sizes, variously-shaped ultrasonic transducer are commercially available；Commercially available miniature ultrasonic transducer units its sizes can be with It is small to only with millimeter calculate magnitude.

Only with regard to miniature ultrasonic transducer units its technique for fixing on general industry application solid body surface itself and Speech, is known general technology for the professional in ultrasonic technology field.This case is not to this expansion superfluous words.

Only with regard to naked PDMS substrates itself micro-channel molding or lithographic technique for, be open-and-shut known technology.

The industrial products market of involved its all size of higher-order of oscillation electric signal transmission cable is on sale.

The structure of the micro fluidic device can also include higher-order of oscillation electric signal generator；The higher-order of oscillation electric signal transmission electricity Cable its other end can be connected with the higher-order of oscillation electric signal generator.

The involved higher-order of oscillation electric signal generator technology of itself, for the professional in ultrasonic technology field, be It is simple and known；The higher-order of oscillation electric signal generator can be customized to ultrasonic instrument specialized factory.

The preferred scope of the miniature ultrasonic transducer units its specified ultrasonic wave transmission powers be between 2 milliwatts and 2000 milliwatts it Between；The preferred scope of the frequency of the miniature ultrasonic transducer units its ultrasonic waves operationally launched be between 100KHz with Between 12MHz.

This case device can further include some annexes, the annex such as multiple tracks electrochemical workstation etc., institute certainly The art-recognized meanings for stating multiple tracks electrochemical workstation are known.Each working electrode for being related in this case microfluidic chip structure and To electrode and reference electrode etc., can respectively via corresponding special the corresponding interface got lines crossed with the multiple tracks electrochemical workstation Coupled.It is described that special to get lines crossed be for each electrode is carried out into phase with each the corresponding interface of the multiple tracks electrochemical workstation The private cable being mutually coupled with.The micro-fluidic chip in this case device, its structure can also include micro-valve, the number of the micro-valve Amount is not limited, and according to actual needs, the micro-valve can be installed in any required position installed in the microfluidic chip structure； For the professional of micro fluidic chip technical field, the art-recognized meanings of itself are known to the word of micro-valve one；The micro-valve The manufacturing technology of itself and the use of technology is also known；The component that the micro-valve is not required.

The diameter of the working electrode can allow to be that any setting is easily installed the suitable diameter for using, it is however recommended to Or say preferred its scope of the diameter between 0.1 micron to 2000 microns；The length of the working electrode can permit Permitted to be that any setting is easily installed the length for using, it is however recommended to or to say the preferred length its scope be at 1 micron To between 15000 microns.

The gold size for being installed in the working electrode surface layer by spraying or point sample instrument point sample or the coating of other appropriate process is quick Sense film, its thicknesses of layers can allow be any setting the sample measuring liquid treated occur electrical signals response thickness, but, push away Preferred thickness is between 10 nanometers and 200 nanometers to the thickness recommended in other words conj.or perhaps.

The cover plate in chip structure, its material can allow to be any electrical insulating property material, for example：Polypropylene, glass Glass, polymethyl methacrylate, dimethyl silicone polymer, etc., in order to make smaller size of micro-fluidic chip, such as do Into the micro-fluidic chip of only 2.0 centimetres to 3.0 centimetres of super-small of length, and realized to ultrasonic wave in the extremely short distance Extremely fast decay, it may be preferred to dimethyl silicone polymer is used as cover plate.Certainly, selected on large-sized micro-fluidic chip It is used as the cover plate using dimethyl silicone polymer, is also that this case technical scheme is allowed.

The preferred scope of the distance between the terminal and the sample introduction end is between 3 centimetres and 10 centimetres.

The cover plate and substrate its thickness can allow be any setting the thickness for being easy to assembling, the thickness of recommendation or say preferably Thickness be between 1.0 millimeters and 5.0 millimeters.Less thickness is conducive to material-saving.

The application method of this case micro-fluidic chip：

This case drives liquid stream to be flowed in the capillary channel of the ten Five-channels micro-fluidic chip with dual drive coupling operating mode It is dynamic, joint-detection is carried out to 15 kinds of tumor markerses respectively using multi-channel electrochemical analyzer device.

The specific detection of this case micro-fluidic chip is as follows using step：

1st, blood serum sample liquid is added in micro-pipe road, in the case where dual drive coupling operating mode drives, various tumor markerses Antigen molecule is by the tumor-marker of the corresponding horseradish peroxidase-labeled of gold size sensitive membrane embedding on electrode surface in each passage Thing specific antibody is captured.

2nd, the tumor markers antigen in the tumor markers specific antibody and blood serum sample of horseradish peroxidase-labeled is formed Immune complex.

3rd, using multi-channel electrochemical analyzer, the electron mediators such as catechol are added, using the above-mentioned reaction of amperometric detection The curent change for causing, is derived from the species and content of various analytes.

4th, result is carried out into comprehensive analysis, comprehensive diagnos is carried out to tumor markers antigen.

It is an advantage of the invention that install miniature ultrasonic in its close position attaching of the terminal of the micro-fluidic chip changing Can device, low-power, the ultrasonic wave of high-frequency band that using the miniature ultrasonic transducer units, it is launched so that without coming to the surface The compatibility learned between modified strong hydrophobic its tube wall of micro-fluidic chip internal pipeline and the test object aqueous solution significantly increases Plus, this is test liquid stream by there is provided a realistic possibility；Meanwhile, using dimethyl silicone polymer substrate its to ultrasound The strong absorbability of ripple, in shorter distance, it is, from the terminal to the only several centimeters the sample introduction end In the very short distance of yardstick, the rapid decrement of ultrasonic intensity is reached, thereby cause institute at the two ends of the micro-fluidic chip The difference of interfacial tension is stated, the difference of the interfacial tension between the two ends can cause a kind of driving force, and this kind is poor because of interfacial tension Its function of driving force is to drive test liquid stream in strong hydrophobic capillary channel originally to the terminal side caused by different To flowing；And the simultaneous Micropump in structure, its function is, leads to inside it in the sample solution and the micro-fluidic chip Interfacial tension between the inwall in road is reduced by the ul-trasonic irradiation, under the increased precondition of alternate compatibility, with the Micropump The two ends that are induced come the ultrasonic wave of mechanicalness pumping strength between interfacial tension difference its driving force for being brought Support mutually, adjust mutually, be mutually coupled, one is pooled in the way of Collaboration and drives test liquid to flow to the terminal side To the strength of flowing；The presence of the Micropump so that the miniature ultrasonic transducer units its ultrasonic wave emissive porwers can be allowed to appropriateness Reduce, this is particularly suitable for for the situation containing ultrasonic wave sensitive composition in detection object；And due to the ultrasonic transducer and its The presence of radiated ultrasonic wave, it is possible to increase alternate compatibility, reduces interfacial tension, and provide two ends interfacial tensions difference its The special driving force brought, then, in this case, test liquid stream its flow resistance in the micro-channel significantly drops Low, correspondingly, the Micropump its running resistance is greatly reduced, and so, the Micropump can just be carried out with than relatively low pumping pressure For the pumping work of the sample solution, because mechanical pumping pressure is greatly reduced, therefore, in such a situation, Just be not susceptible to because sample introduction terminal tool pumping pressure it is excessive caused by the sample introduction end and its near zone the micro-channel drum Between bubble, expansion, deformation, distortion and the region substrate and cover plate the problems such as stripping etc.；This case dual drive coupling running Scheme and increased for the sample fluid flow action it is handling, can allow for use ultrasonic intensity, ultrasonic frequency, Micropump pump power, Micropump pumping pressure etc. multiple indexs include flowing forward to be directed to the flow rate of the flowing, flowing and act Dynamic or pause flowing or acceleration flowing etc. flowing action carries out multi-parameter and accurately manipulates；By this case dual drive coupling running side Case, repaiies entirely without that must carry out any surface chemistry to the substrate of the dimethyl silicone polymer material its micro-channel etc. relevant surfaces Decorations or surface chemical modification, have altogether dispensed with the laborious procedures of the surface chemical modification or surface chemical modification；On the other hand, should The ultrasonic wave of low-power, high-frequency band, additionally it is possible to contain the large biological molecule in sample in the literalness naked poly dimethyl silicon Absorption on oxyalkyl piece its inner surface of pipeline, and then contain the dimethyl silicone polymer substrate its body phase to the large biological molecule The effect of swallowing up；The Reversible binding thing of the antigen, antibody and antigen and antibody is all belonging to described large biological molecule certainly Type；Because described suction-operated and the described effect of swallowing up effectively are contained, therefore, dependence test result will more Plus can objectively reflect actual conditions；The effect of the low-power, high-frequency band ultrasonic wave, also includes facilitating antigen, resists certainly The quick of Reversible binding reaction between body is reached, and this causes that dependence test is operated can complete with than speed faster.

As described above, the presence of the Micropump so that the miniature ultrasonic transducer units its ultrasonic wave emissive porwers can be allowed to fit Degree is reduced, then, the feature helps to protect the working electrode its sensitive coating, is allowed to from ultrasound injury.

Based on this case scheme, repaiied completely without the surface chemistry carried out for the dimethyl silicone polymer substrate its relevant surfaces Decorations or surface chemical modification operation, therefore, this surface chemical modification layer or surface chemical modification layer not need presence, So, its body phase interior polymer molecule of the dimethyl silicone polymer substrate is constantly diffused to the surface, migrated caused by it automatically Damaging influence to surface chemical modification layer or surface chemical modification layer does not just exist yet.

The technical scheme of this case has dissolved its application related one to dimethyl silicone polymer substrate addressed above totally Row technical barrier.Based on this case scheme, the polydimethyl siloxane material of this kind very cheap and easily processing and fabricating just has can Can in the micro-fluidic chip its prepare, production, using etc. field play bigger effect.

The framework of this case micro-fluidic chip its large-scale integrated, determines it among actual test application, to Serum samples The need for measure smaller, this body and mind for helping to reduce related subject is damaged.

Brief description of the drawings

Fig. 1 is its rough outside side view of this case device.

In figure, 1 is the substrate of dimethyl silicone polymer material, and 2 is cover plate, and 3 is higher-order of oscillation electric signal transmission cable, 4 It is miniature ultrasonic transducer units, 5 is the sample introduction end of the micro-fluidic chip, and 6 is the terminal of the micro-fluidic chip, 7 It is Micropump, 8 is infusion tube；Arrow in legend indicate the micro-fluidic chip its in actual motion, by pressure at two ends Difference drives, the flow direction of its test liquid stream.

Specific embodiment

In this case that Fig. 1 is shown embodiment, the example is characterized in, the structure of the device includes micro-fluidic chip, should The structure of micro-fluidic chip includes installing bonded to each other substrate 1 and cover plate 2 together, and the substrate 1 and cover plate 2 are plate Contain the conduit formed via mould pressing process or etching technics in shape thing or tablet, that face towards the cover plate 2 of the substrate 1 Structure, substrate 1 being installed together bonded to each other has been built into the micro-fluidic chip containing pipeline configuration jointly with the cover plate 2, The locations of structures of the pipeline is located at the substrate 1 and the cover plate 2 juncture area bonded to each other, and the two ends of the pipeline are micro- with this respectively The sample introduction end 5 of fluidic chip and terminal 6 are connected, and the sample introduction end 5 is the injection end of the micro-fluidic chip sample solution, the end End 6 is the terminal of sample solution flowing in its chip when the actual sample introduction of the micro-fluidic chip is tested, the terminal 6 and the sample introduction end 5 It is located remotely from each other, the distance between the terminal 6 and the sample introduction end 5 are different in the pipeline between 3 centimetres and 10 centimetres On position in order or backward is equiped with working electrode and to electrode and reference electrode, the order refers to the reference electricity Extremely locations of structures closer to the position of the terminal 6, the backward refer to the reference electrode locations of structures closer to it is described enter The position of sample end 5, the working electrode is resisted by conductive electrode and the tumor markers that embedded being attached on the conductive electrode The gold size sensitive membrane of body is constituted, and the construction of the pipeline is presented parallel construction, and the pipeline in parallel construction is by 15 branched pipes Road is in parallel to be constituted, described that the profile that the pipeline of parallel construction its appearance profile is similar to parallel circuit, the work electricity is presented The quantity of pole is 15, and 15 installation positions of working electrode are located in 15 laterals respectively, and, Tumor markers antibody in the sensitive membrane structure of its top layer gold size of 15 working electrodes is respectively to tumor markers antigen energy Specific binding 15 kinds of tumor markers antibody materials, 15 kinds of antibody materials be respectively tumor markers antibody A FP, CEA、CA242、CA125、CA199、CA153、CA724、CA50、NSE、CYFRA21-1、FPSA、TPSA、β-hCG、 SCCA and β 2-MG, the antigen is the antigen of broad sense, and the antibody is the antibody of broad sense, the working electrode its material It is argent material, gold material, carbon material or thermal decomposition conducting polymer material, the working electrode its pattern presentation post Shape, sheet or thread, the substrate 1 its material is dimethyl silicone polymer material, and its surface of substrate 1 is the table of primary form Face, its material for being intended to refer to not by any surface chemical modification or any surface chemical modification of the surface of the primary form The surface of the primary form of matter, the structure of the device also includes miniature ultrasonic transducer units 4, and, higher-order of oscillation electric signal is passed Transmission cable 3, one end of the higher-order of oscillation electric signal transmission cable 3 links together with the miniature ultrasonic transducer units 4, and this is micro- Type ultrasonic transducer 4 is installed in the position of the neighbouring described terminal 6 of the cover plate 2 or substrate 1 of the micro-fluidic chip with attaching； The miniature ultrasonic transducer units 4 its major functions is the ultrasonic wave launched using it when the actual sample introduction of micro-fluidic chip is tested To reduce the interfacial tension between sample solution and the inwall of its inner passage of micro-fluidic chip, can be compatible, also, Using the distance between the sample introduction end 5 and the terminal 6 and the installation position of miniature ultrasonic transducer units 4 difference and its The difference on ultrasonic intensity experienced, its interfacial tension of sample introduction end 5 described in induced synthesis and its interface of the terminal 6 Difference between power, the interfacial tension difference between the micro-fluidic chip two ends 5,6 can be at the two ends of the micro-fluidic chip 5th, pressure gap is formed between 6, the pressure gap can drive sample solution to be flowed to the direction of the terminal 6；The miniature ultrasonic Wave transducer 4 its function also include the ultrasonic wave launched with it check contained large biological molecule in sample its in the miniflow Absorption on control chip its inner passage inner surface, and then check the substrate 1 of the dimethyl silicone polymer material its body phase to the life The effect of swallowing up of thing macromolecular；It is soft and have the substrate 1 of the dimethyl silicone polymer material of elasticity its function and include with it to super The property that sound wave absorbs strongly, is absorbed strongly to ultrasonic wave, and thereby in the micro-fluidic chip, the terminal 6 arrives the sample introduction end The rapid decrement of ultrasonic intensity is realized within limited short distance between 5；And, Micropump 7, the Micropump 7 and the sample introduction End 5 connects；The function of the Micropump 7 is, the interface between the inwall of the sample solution and its inner passage of micro-fluidic chip Tension force is reduced by the ul-trasonic irradiation, under the increased precondition of alternate compatibility, strength is pumped with the mechanicalness of the Micropump 7 Supported mutually come interfacial tension difference its driving force brought between the two ends 5,6 of ultrasonic wave induction, mutually Mutually adjust, be mutually coupled, one is pooled in the way of Collaboration and drives test liquid to flow to the power of the direction of the terminal 6 flowing Amount.

Arrow in legend indicate the micro-fluidic chip its in actual motion, by two ends 5,6 pressure differentials drive, its examination The flow direction of sample liquid stream.

Fig. 1 is depicted without the associate members such as the higher-order of oscillation electric signal generator.

Involved miniature ultrasonic transducer units 4 are commercially available；Can also be customized to ultrasonic transducer producer.

Involved higher-order of oscillation electric signal transmission cable 3 is commercially available；Can also be customized to ultrasonic transducer producer.

Involved higher-order of oscillation electric signal generator market has the product close to needs commercially available；Can also be customized to relevant manufacturers.

Micropump 7 involved by legend can be customized to specialized factory.

Its inner passage of the micro-fluidic chip involved by this case is the pipeline of hydrophobic capillary form.

The Micropump both can be the Micropump of external form；The Micropump can also be done into or say inside the embedded micro-fluidic chip The Micropump of the built in version of its sample introduction end structure position or the sample introduction end Near-neighbor Structure position.Micropump in legend is external form Micropump；The Micropump can certainly do into or say embedded microfluidic chip structure inside sample introduction end or its Near-neighbor Structure position The Micropump of the built in version, its basic structure key element is identical with the Micropump of external form.

Each working electrode in this example structure and electrode and reference electrode via each special cable or can be said respectively Get lines crossed respectively with the corresponding cable interface of the multiple tracks electrochemical workstation as annex or say interface of getting lines crossed and couple.

In view of its form of this case related text expresses that it is described above pipeline of the presentation parallel construction is clear enough It is clear, the concrete form of the pipeline in this kind of micro-fluidic chip of this case is no longer specifically illustrating in this case embodiment.

Antibody described in this case refers to the antibody of broad sense；Antigen described in this case refers to the antigen of broad sense；It is immunized described in this case multiple Compound refers to the immune complex of broad sense.

Claims (10)

1. using the chip apparatus for joint inspection Diagnostic Value of Several Serum Tumor Markers of hydrophobic substrate, it is characterised in that the structure bag of the device Micro-fluidic chip is included, the structure of the micro-fluidic chip includes installing bonded to each other substrate and cover plate together, the substrate and lid Piece is plate object or tablet, and what is formed via mould pressing process or etching technics contained in that face towards the cover plate of the substrate Channel structure, substrate being installed together bonded to each other has been built into the micro-fluidic core containing pipeline configuration jointly with the cover plate Piece, the locations of structures of the pipeline is located at the substrate and cover plate juncture area bonded to each other, and the two ends of the pipeline are micro- with this respectively The sample introduction end of fluidic chip and terminal are connected, and the sample introduction end is the injection end of the micro-fluidic chip sample solution, and the terminal is this The terminal that sample solution flows in its chip when the actual sample introduction of micro-fluidic chip is tested, the terminal is located remotely from each other with the sample introduction end, should The distance between terminal and the sample introduction end are in order or inverse on diverse location in the pipeline between 3 centimetres and 10 centimetres Sequence is equiped with working electrode and to electrode and reference electrode, and the order refers to the reference electrode its locations of structures and more leans on The nearly terminal location, the backward refers to the reference electrode locations of structures closer to the sample introduction end position, the work Electrode is made up of conductive electrode and the gold size sensitive membrane for having embedded tumor markers antibody being attached on the conductive electrode, The construction of the pipeline is presented parallel construction, and the pipeline in parallel construction is made up of 15 lateral parallel connections, the presentation The pipeline of parallel construction its appearance profile is similar to the profile of parallel circuit, and the quantity of the working electrode is 15, should 15 installation positions of working electrode are located in 15 laterals respectively, and, 15 working electrode its tables Tumor markers antibody in layer gold size sensitivity membrane structure is respectively that 15 kinds can specifically bound to tumor markers antigen swell Tumor markers antibody materials, 15 kinds of antibody materials be respectively tumor markers antibody A FP, CEA, CA242, CA125, CA199, CA153, CA724, CA50, NSE, CYFRA21-1, FPSA, TPSA, β-hCG, SCCA and β 2-MG, it is described anti- Original is the antigen of broad sense, and the antibody is the antibody of broad sense, the working electrode its material be argent material, gold material, Carbon material or thermal decomposition conducting polymer material, the working electrode its pattern are presented column, sheet or thread, the substrate its Material is dimethyl silicone polymer material, and its surface of the substrate is the surface of primary form, the surface of the primary form its be meant to Be not by any surface chemical modification or the surface of the primary form of the material of any surface chemical modification, the device Structure also includes miniature ultrasonic transducer units, and, higher-order of oscillation electric signal transmission cable, higher-order of oscillation electric signal transmission electricity One end of cable links together with the miniature ultrasonic transducer units, and the miniature ultrasonic transducer units are installed in the micro-fluidic core with attaching The position of the cover plate of piece or the neighbouring described terminal of substrate；The miniature ultrasonic transducer units its major functions is in micro-fluidic chip reality When border sample introduction is tested, the ultrasonic wave launched using it reduce sample solution and its inner passage of micro-fluidic chip inwall it Between interfacial tension, can be compatible, also, using the sample introduction end and the terminal and the miniature ultrasonic transducer units Difference in the distance between installation position difference and its ultrasonic intensity experienced, its boundary of sample introduction end described in induced synthesis Difference between face tension force and the terminal its interfacial tension, the interfacial tension difference between the micro-fluidic chip two ends can be at this Pressure gap is formed between the two ends of micro-fluidic chip, the pressure gap can drive sample solution to the end flow；This is micro- Type ultrasonic transducer its function also include the ultrasonic wave launched with it check contained large biological molecule in sample its at this Absorption on micro-fluidic chip its inner passage inner surface, and then check the substrate of the dimethyl silicone polymer material its body phase to this The effect of swallowing up of large biological molecule；It is soft and have the substrate of the dimethyl silicone polymer material of elasticity its function and include with it to super Sound wave strongly absorb property, ultrasonic wave is absorbed strongly, and thereby micro-fluidic chip terminal to the sample introduction end it Between limited short distance within realize the rapid decrement of ultrasonic intensity；And, Micropump, the Micropump is connected with the sample introduction end； The function of the Micropump is that the interfacial tension between the inwall of the sample solution and its inner passage of micro-fluidic chip receives the ultrasound Ripple acts on and reduces, under the increased precondition of alternate compatibility, pumps strength with the mechanicalness of the Micropump to be lured with the ultrasonic wave Interfacial tension difference its driving force brought between the two ends led is supported mutually, adjusts mutually, is mutually coupled, with The mode of Collaboration pools one and drives test liquid to flow to the strength of the terminal direction flowing.

2. the chip apparatus for joint inspection Diagnostic Value of Several Serum Tumor Markers using hydrophobic substrate according to claim 1, its feature It is that the pipeline is capillary channel including the lateral.

3. the chip apparatus for joint inspection Diagnostic Value of Several Serum Tumor Markers using hydrophobic substrate according to claim 1, its feature It is that the thermal decomposition conducting polymer is the conductive material formed after being heat-treated through anoxybiotic by polyimides or polyacrylonitrile.

4. the chip apparatus for joint inspection Diagnostic Value of Several Serum Tumor Markers using hydrophobic substrate according to claim 1, its feature Be, the width or diameter of the working electrode between 0.1 micron to 2000 microns, and, the working electrode Length is between 1 micron to 15000 microns.

5. the chip apparatus for joint inspection Diagnostic Value of Several Serum Tumor Markers using hydrophobic substrate according to claim 1, its feature It is that the thickness of the gold size sensitive membrane is between 10 nanometers and 200 nanometers.

6. the chip apparatus for joint inspection Diagnostic Value of Several Serum Tumor Markers using hydrophobic substrate according to claim 1, its feature It is that its material of the cover plate in structure is dimethyl silicone polymer material.

7. the chip apparatus for joint inspection Diagnostic Value of Several Serum Tumor Markers using hydrophobic substrate according to claim 1, its feature It is that the Micropump is miniature piezoelectric pump, miniature peristaltic pump or miniature air driven pump.

8. the chip apparatus for joint inspection Diagnostic Value of Several Serum Tumor Markers using hydrophobic substrate according to claim 1, its feature It is that the cover plate in structure and substrate its thickness are between 1.0 millimeters and 5.0 millimeters.

9. the chip apparatus for joint inspection Diagnostic Value of Several Serum Tumor Markers using hydrophobic substrate according to claim 1, its feature Be that the structure of the micro fluidic device also includes higher-order of oscillation electric signal generator, the higher-order of oscillation electric signal transmission cable its The other end is connected with the higher-order of oscillation electric signal generator.

10. the chip apparatus for joint inspection Diagnostic Value of Several Serum Tumor Markers using hydrophobic substrate according to claim 1, its feature It is that the miniature ultrasonic transducer units its specified ultrasonic wave transmission powers is between 2 milliwatts and 2000 milliwatts, and this is miniature super The frequency of acoustic wave transducer its ultrasonic wave operationally launched is between 100KHz and 12MHz.