KR20070106877A - A batteryless fluid transfering lab-on-a-chip for portable dianostics - Google Patents

A batteryless fluid transfering lab-on-a-chip for portable dianostics Download PDF

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KR20070106877A
KR20070106877A KR1020060039260A KR20060039260A KR20070106877A KR 20070106877 A KR20070106877 A KR 20070106877A KR 1020060039260 A KR1020060039260 A KR 1020060039260A KR 20060039260 A KR20060039260 A KR 20060039260A KR 20070106877 A KR20070106877 A KR 20070106877A
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South Korea
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chip
lab
substrate
reaction chamber
buffer
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KR1020060039260A
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Korean (ko)
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양상식
박신욱
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아주대학교산학협력단
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    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
    • B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61B—DIAGNOSIS; SURGERY; IDENTIFICATION
    • A61B5/00—Measuring for diagnostic purposes; Identification of persons
    • A61B5/145—Measuring characteristics of blood in vivo, e.g. gas concentration, pH value; Measuring characteristics of body fluids or tissues, e.g. interstitial fluid, cerebral tissue
    • A61B5/1486—Measuring characteristics of blood in vivo, e.g. gas concentration, pH value; Measuring characteristics of body fluids or tissues, e.g. interstitial fluid, cerebral tissue using enzyme electrodes, e.g. with immobilised oxidase
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
    • G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
    • G01N27/28—Electrolytic cell components
    • G01N27/30—Electrodes, e.g. test electrodes; Half-cells
    • G01N27/327—Biochemical electrodes, e.g. electrical or mechanical details for in vitro measurements
    • G01N27/3271—Amperometric enzyme electrodes for analytes in body fluids, e.g. glucose in blood
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/5302—Apparatus specially adapted for immunological test procedures
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G01N35/00029—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor provided with flat sample substrates, e.g. slides
    • G01N2035/00099—Characterised by type of test elements
    • G01N2035/00158—Elements containing microarrays, i.e. "biochip"

Abstract

A battery-less fluid transferring lab-on-a-chip for portable diagnosis is provided to perform the separation, recognition, and signal detection of a vital molecule effectively. A method for manufacturing a battery-less fluid transferring lab-on-a-chip for portable diagnosis includes the steps of: applying a micro machining process for a lower substrate of glass or polymer to form a sensing electrode on a lower substrate; forming a fluid passage(8), a reaction chamber(3), a waste chamber(9), and a minute hole(4) for connecting upper and lower portions on an intermediate substrate using a polymer minute processing method; forming a storage passage, an entrance, and a waste outlet on an upper substrate; and adhering the respective substrates to each other using a plasma surface process.

Description

무전원 유체 이송 방식 휴대 진단용 랩온어칩 { A batteryless fluid transfering lab-on-a-chip for portable dianostics}A batteryless fluid transfering lab-on-a-chip for portable dianostics}

도 1은 본 발명에 따른 무전원 유체 이송 방식 휴대 진단용 랩온어칩의 평면도.1 is a plan view of a non-powered fluid transfer type portable diagnostic lab-on-a-chip according to the present invention.

도 2는 본 발명에 따른 무전원 유체 이송 방식 휴대 진단용 랩온어칩의 분해도.Figure 2 is an exploded view of a non-powered fluid transfer method portable diagnostic lab-on-a-chip according to the present invention.

도 3은 본 발명에 따른 무전원 유체 이송 방식 휴대 진단용 랩온어칩의 동작도.Figure 3 is an operation of a non-powered fluid transfer method portable diagnostic lab-on-a-chip according to the present invention.

<도면의 주요부분에 대한 부호의 설명><Description of the symbols for the main parts of the drawings>

1 : 시료 주입구 2 : 시료 미세 유로 1 sample injection hole 2 sample microchannel

3 : 반응 챔버 4 : 상ㆍ하부 연결 미세 구멍 3: reaction chamber 4: upper and lower connection micropores

5 : 센싱 전극 6 : 유체 정지 구간 5: sensing electrode 6: fluid stop section

7 : 완충액 주입구 8 : 완충액 저장 유로  7: buffer inlet port 8: buffer storage flow path

9 : 폐기 챔버 10 : 폐기 출구 9: waste chamber 10: waste outlet

11 : 공기저장 챔버 12 : 하부 기판11 air storage chamber 12 lower substrate

13 : 중간 기판 14 : 상부 기판13: intermediate substrate 14: upper substrate

본 발명은 휴대 진단용 면역 분석 시스템 실용화 분야에 적용 가능한 무전원 유체 이송 방식 휴대 진단용 랩온어칩에 관한 것으로, 더욱 상세하게는 시료(혈액)가 주입되면 모세관 인력에 의해 칩 안에 내장되어 있는 면역 센서 영역으로 자동적으로 이송되며, 일정한 시간 경과 후 공기압에 의한 자가 세척을 할 수 있는 유체 제어 시스템과 항원 또는 항체의 존재 유무 및 농도를 효율적으로 검출하는 통합형 바이오센서가 내장된 바이오칩과 이를 제작하는 방법에 관한 것이다. 최근 의료 및 생명 분야에서 소량의 혈액이나 체액만으로 단백질, DNA, 면역반응, 세포 등에서 얻어지는 생체정보를 총체적으로 감지하여 건강 상태와 질병을 진단할 수 있는 휴대용 진단기에 대한 연구가 주목받고 있다. 하지만 기존의 면역 분석시스템에는 시료의 양이 많이 필요하며, 시료를 제어하기 위해 부가적인 전원 장치가 필요하기 때문에 소형화에 큰 어려움이 있다. 휴대가능한 진단 시스템을 구축하기 위해 개발되어진 바이오센서의 고감도 특성을 유지하면서, 기존의 분석 도구와 차별성을 갖는 제품의 소형화, 편의성, 정확성, 신뢰성을 가져야 한다. 랩온어칩과 초소형 종합 분석 시스템과 같이 미세 유체 소자를 사용한 분석 시스템의 경우 분석 장비사 소형화되어 필요한 시료의 양이 작아지게 되며, 시료 주입 이후 대부분의 검사공정이 자동으로 진행되어 필요 인력 및 분석 시간이 줄어든다. 또한 효율적인 생화학 재료 사용 및 표면 처리 공정을 통해 주요 구성 요소를 단순화 시켜 제조비용을 절감시킬 수 있는 공정 기술이 요구된다.The present invention relates to a non-powered fluid transfer type portable diagnostic lab-on-a-chip applicable to the practical application of the immunoassay system for portable diagnostics. More specifically, the present invention relates to an immune sensor region embedded in a chip by capillary attraction when a sample (blood) is injected. The present invention relates to a biochip equipped with a fluid control system which is automatically transferred and can be self-cleaned by air pressure after a predetermined time, and an integrated biosensor which efficiently detects the presence and concentration of antigen or antibody, and a method of manufacturing the same. . Recently, the medical and life fields have attracted attention for a portable diagnostic device that can detect health information and diseases by collectively detecting biological information obtained from proteins, DNA, immune responses, and cells with only a small amount of blood or body fluids. However, the existing immunoassay system requires a large amount of sample, and there is a great difficulty in miniaturization because an additional power supply is required to control the sample. While maintaining the high sensitivity of the biosensor developed to build a portable diagnostic system, it should have the miniaturization, convenience, accuracy and reliability of the product which is different from the existing analysis tools. In the case of analysis systems using microfluidic devices such as lab-on-a-chip and ultra-compact comprehensive analysis systems, the size of analytical equipment can be miniaturized, and the amount of sample required is reduced. This decreases. There is also a need for process technologies that can reduce manufacturing costs by simplifying key components through efficient biochemical material use and surface treatment processes.

발명에서 제시한 면역센싱 랩온어칩은 상기와 같은 문제점을 해소하고 효율을 극대화하기 위해 마이크로머시닝 기술과 초소형 유체 제어 기술을 적용하여 수 마이크로(10-6) 리터의 시료를 별도의 외부 전원 없이 모세관 인력과 구간 표면처리만을 이용하여 구조적으로 제어한다. 랩온어칩 안에 있는 센싱 전극에서 생체분자 간 인식반응 및 생체촉매반응을 전기화학적인 방식으로 신호를 검출하여 측정시스템의 일체화, 모듈화가 가능하다. 본 발명의 랩온어칩에서 완충액을 이송 제어하기위해 공기 챔버 안에 있는 공기를 특별한 외부 장비 없이 손으로 압축하여 압축된 공기의 부피만큼 완충액을 밀어내게 되는 간단한 원리를 적용하였다. 위에서 상기된 모든 구조가 마이크로머시닝 기술로 제작하기 때문에 제작 공정이 간단하며 집적화가 가능하다.Immune sensing lab-on-a-chip in the present invention, the number of micro-capillary (10-6) l of the sample by applying a micro-machining technology and compact fluid control technology to solve the above problems and to maximize efficiency without the need for external power Structural control using only manpower and section surface treatment. The sensing electrodes in the lab-on-a-chip can detect signals between biomolecules and biocatalytic reactions in an electrochemical manner to integrate and measure the measurement system. In order to transfer and control the buffer in the lab-on-a-chip of the present invention, a simple principle of compressing the air in the air chamber by hand without special external equipment and pushing the buffer by the volume of the compressed air was applied. Since all the structures described above are manufactured by micromachining technology, the manufacturing process is simple and integration is possible.

본 발명은 유리 또는 생체 적합한 폴리머 재질의 하부 기판에 마이크로머시닝 기술을 적용하여 마이크로 센싱 전극을 제작하는 단계, 폴리머 재질의 중간 기판에 플라스틱 가공 공정을 이용하여 시료의 입구와 미세 유로, 반응 챔버, 폐기 챔버 및 미세 공기구멍, 상부기판과 연결되는 미세 구멍을 제작하는 단계, 폴리머 재질의 상부 기판에 완충액을 저장할 저장 유로, 완충액을 밀어낼 수 있는 공기 챔버 및 초미세 공기 유로를 제작하는 단계, 각 기판을 접합하는 단계로 이루어진다. 상기 단계들로 제작된 랩온어칩은 도3(가)와 같이 완충액이 완충액 주입구에서 상하부연결 구멍 앞까지 완충액 저장 유로에 주입되는 단계, 도3(나)와 같이 시료가 시료주입구(1)에서 모세관 인력에 의해 자동적으로 반응 챔버까지 이송되어 항체가 부착되어 있는 센서에 착상되는 단계, 도3(다)와 같이 손으로 눌려진 공압에 의해 반응 챔버에 존재하던 시료를 폐기 챔버로 밀어내고 효소가 포함된 완충액으로 효소 산화시키는 단계, 순환전압전류법에 의해 산화된 효소의 신호를 검출하는 단계로 이루어진다.The present invention is a step of manufacturing a micro-sensing electrode by applying micromachining technology to the lower substrate of glass or biocompatible polymer material, using the plastic processing process on the intermediate substrate of the polymer material, the inlet and the micro flow path of the sample, reaction chamber, disposal Manufacturing a chamber and a micro air hole, a micro hole connected to the upper substrate, a storage flow path for storing the buffer solution in a polymer upper substrate, an air chamber capable of pushing the buffer solution, and an ultra fine air flow path, each substrate It consists of a step of bonding. The lab-on-a-chip manufactured by the above steps is a step in which the buffer is injected into the buffer storage passage from the buffer inlet to the upper and lower connection holes as shown in FIG. 3 (a), and the sample is injected from the sample inlet 1 as shown in FIG. Automatically transferred to the reaction chamber by capillary attraction and implanted on the sensor to which the antibody is attached. As shown in FIG. 3 (c), the sample existing in the reaction chamber is pushed into the waste chamber by the pneumatic pressure, and the enzyme is contained. And oxidizing the enzyme with the prepared buffer, and detecting the signal of the oxidized enzyme by cyclic voltammetry.

이하 첨부된 도면에 의해 상세히 설명하면 다음과 같다.Hereinafter, described in detail by the accompanying drawings as follows.

도 1과 2는 각각 본 발명의 무전원으로 구동하는 휴대 진단용 랩온어칩의 평면도와 분해도이다. 제안된 랩온어칩은 생체면역 반응 챔버(3)와 4개의 센싱 전극(5), 시료의 이동을 제어하는 유로(2)와 완충액을 저장하는 유로(8), 공기 저장 챔버(11), 폐기 챔버(9)로 구성되어 있다. 하부 기판(12)은 생체면역 센싱 전극(5)과 시료를 정지하기 위한 유체 정지 구간(6)을 가지고 있다. 중간 기판(13)은 시료가 흘러갈 수 있는 미세 채널(2)과 센싱 전극으로 흘러갈 수 있는 반응 챔버(3)을 갖고 있으며 완충액이 반응 챔버로 흘러 들어갈 수 있는 상하부 미세구멍(4), 반응이 끝난 시료가 완충액에 의해 밀려 나가게 되는 폐기 챔버(9)로 이루어져 있다. 상부 기판(14)은 각각의 시료와 완충액을 위한 입구(1, 7)와 폐기 출구(10), 완충액을 저장하는 유로(8), 완충액을 반응 챔버로 밀어내는 공압을 위한 공기 저장 챔버(11)가 있다. 최종적으로 무전원 유체 이송 방식 휴대 진단용 랩온어칩은 상부 기판(14)과 중간 기판(13) 하부 기판(12)이 접합된 구조로 되어있다.1 and 2 are a plan view and an exploded view, respectively, of the portable diagnostic lab-on-a-chip driven by the power supply of the present invention. The proposed lab-on-a-chip consists of a bioimmune reaction chamber (3) and four sensing electrodes (5), a flow path (2) for controlling the movement of the sample, a flow path (8) for storing the buffer, an air storage chamber (11), and disposal. It consists of the chamber 9. The lower substrate 12 has a bioimmune sensing electrode 5 and a fluid stop section 6 for stopping the sample. The intermediate substrate 13 has a microchannel 2 through which a sample can flow and a reaction chamber 3 through which a sensing electrode can flow, and an upper and lower micropores 4 through which a buffer can flow into the reaction chamber. This finished sample consists of a waste chamber 9 which is pushed out by the buffer solution. The upper substrate 14 includes inlets 1 and 7 for each sample and buffer and a waste outlet 10, a flow path 8 for storing the buffer, and an air storage chamber 11 for pneumatic pressure to push the buffer into the reaction chamber. There is). Finally, the non-powered fluid transfer type portable diagnostic lab-on-a-chip has a structure in which the upper substrate 14 and the intermediate substrate 13 and the lower substrate 12 are bonded to each other.

무전원 유체 이송 방식 휴대 진단용 랩온어칩의 제작 공정은 다음과 같다. 하부 기판(12)에 화학 기상 증착 방식으로 크롬과 금을 증착한 후, 마이크로 머시닝 기술을 이용하여 센싱 전극(5)과 유체정지구간(6)을 제작한다. 중간 기판(13)을 제작하기 위해 실리콘에 음성 후막 감광제를 두껍게 도포한 후 사진 식각 기술을 이용하여 반응 챔버, 시료 이송 유로, 폐기 챔버를 패턴을 형성하는 틀을 제작한다. 제작된 틀 위에 액체 폴리머를 원하는 두께만큼 도포하여 진공 오븐을 이용하여 폴리머 안에 있는 미세 공기들을 제거한 후 굳힌다. 굳어진 폴리머를 틀에서 때어낸 후 미세 기계 가공 방법을 이용하여 상하부를 연결하는 미세 구멍을 형성하여 중간 기판을 제작한다. 중간 기판과 동일한 제작 방법을 이용하여 완충액 저장 유로, 공기 저장 챔버를 가지는 상부 기판을 제작한다. 제작된 각각의 기판을 산소 플라즈마 표면 처리를 한 후 접합하여 무전원 유체 이송 방식 휴대 진단용 랩온어칩을 완성한다.The fabrication process of the portable diagnostic lab-on-a-chip for non-powered fluid transfer is as follows. After depositing chromium and gold on the lower substrate 12 by chemical vapor deposition, the sensing electrode 5 and the fluid stop section 6 are manufactured by using micromachining technology. In order to fabricate the intermediate substrate 13, a thick thick film photosensitive agent is applied to silicon, and then a frame is formed to form patterns of the reaction chamber, the sample transfer channel, and the waste chamber by using photolithography. Apply the liquid polymer to the desired thickness on the mold, remove the fine air in the polymer using a vacuum oven and harden. After the hardened polymer is removed from the mold, an intermediate substrate is manufactured by forming fine holes connecting the upper and lower parts by using a micromachining method. An upper substrate having a buffer storage flow path and an air storage chamber was fabricated using the same fabrication method as the intermediate substrate. Each fabricated substrate is bonded after oxygen plasma surface treatment to complete a lab-on-a-chip for non-powered fluid transfer.

이상에서 상술한 바와 같이 본 발명에서 제안한 무전원 유체 이송 방식 휴대 진단용 랩온어칩은 마이크로머시닝 기술로 제작하여 초소형화가 가능하고 생체 분자의 분리, 인식, 신호 검출이 단일 칩 안에서 효과적으로 수행된다. 본 발명의 초소형 랩온어칩은 소량의 혈액이나 체액만으로도 단백질, DNA, 면역반응, 세포 등에서 얻어지는 생체 정보를 총체적으로 감지하여 건강상태와 질병을 진단할 수 있는 시스템을 구현할 수 있고, 향후 질병 진단용 생체 마이크로 시스템에 의한 다중 질병진단을 통하여 의료기분야에서의 직접적인 응용이 가능하다. 또, 대량생산을 통한 생산 단가 절감으로 대중화 및 보급에도 크게 기여함으로써 기초과학과 산업의 발전 및 국민의 복지에도 직접 혹은 간접적으로 이바지할 것으로 기대된다.As described above, the non-powered fluid transfer type portable diagnostic lab-on-a-chip proposed in the present invention can be miniaturized by micromachining technology, and the separation, recognition, and signal detection of biomolecules are effectively performed in a single chip. The ultra-small lab-on-a-chip of the present invention can implement a system capable of diagnosing a health condition and a disease by collectively detecting biological information obtained from proteins, DNA, immune responses, cells, etc., even with a small amount of blood or body fluids, and for future disease diagnosis. Multiple disease diagnosis by micro system enables direct application in medical field. In addition, it is expected to contribute directly or indirectly to the development of basic science and industry and the welfare of the people by greatly contributing to popularization and dissemination through the reduction of production cost through mass production.

Claims (2)

무전원 유체 이송 방식 휴대 진단용 랩온어칩 제작 방법에 있어서,In the non-power fluid transfer method portable diagnostic lab-on-a-chip manufacturing method, 유리 또는 폴리머 재질의 하부 기판에 마이크로머시닝 공정을 적용하여 센싱 전극과 , 유체정지 구간을 형성하는 단계,Forming a sensing electrode and a fluid stop section by applying a micromachining process to the lower substrate of glass or polymer material, 중간 기판에 폴리머 미세 가공 방법을 이용하여 유체 유로, 반응 챔버 및 폐기 챔버, 기계 또는 레이저 미세 가공을 이용한 상하부 연결 미세 구멍을 제작하는 단계,Fabricating upper and lower connecting micropores in the intermediate substrate using a polymer micromachining method using a fluid microchannel, a reaction chamber and a waste chamber, a machine or laser micromachining, 상부 기판에 폴리머 미세 가공 방법을 이용하여 완충액을 저장할 수 있는 저장 유로와 기계 또는 레이저 미세 가공을 이용한 각각의 유체를 주입할 수 있는 입구, 폐기 출구를 제작하는 하는 단계 Manufacturing a storage flow path for storing a buffer solution using a polymer micromachining method, and an inlet and a waste outlet for injecting respective fluids using a mechanical or laser micromachining process on an upper substrate; 플라즈마 표면 처리를 이용하여 각각의 기판을 접합하는 단계Bonding each substrate using plasma surface treatment 상기의 모든 항을 포함하여 제작됨을 특징으로 하는 무전원 유체 이송 방식 휴대 진단용 랩온어칩을 제작하는 방법 Method for manufacturing a non-powered fluid transfer method mobile diagnostic lab-on-a-chip characterized in that it is produced including all the above terms 상기 단계들로 제작된 무전원 유체 이송 방식 휴대 진단용 랩온어칩의 작동 방법에 있어서,In the method of operating a non-powered fluid transfer method portable diagnostic lab-on-a-chip manufactured by the above steps, 모세관 인력을 이용하여 시료를 반응 챔버에 충전시키는 단계, Filling the reaction chamber with a sample using capillary attraction; 랩온어칩 안의 반응 챔버에서 착상 단계 및 짧은 시간 동안 전극표면과 반응 하는 단계,In the reaction chamber in the lab-on-a-chip and reacting with the electrode surface for a short time, 공기압을 이용하여 완충액을 반응 챔버로 이송하는 단계,Transferring the buffer to the reaction chamber using air pressure, 미세 유로를 통해 이송된 소량의 완충액을 통해 전기화학적 신호를 검출하는 단계, 상기 순서를 단일 칩 안에서 구현하도록 작동되는 방식.Detecting the electrochemical signal through a small amount of buffer transferred through the microchannel, the manner in which the sequence is operated to implement within a single chip.
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