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WO2023065645A1 - Puce microfluidique d'homogénéisation de concentration à base d'électro-mouillage et procédé d'homogénéisation de concentration - Google Patents

Puce microfluidique d'homogénéisation de concentration à base d'électro-mouillage et procédé d'homogénéisation de concentration Download PDF

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Publication number
WO2023065645A1
WO2023065645A1 PCT/CN2022/091636 CN2022091636W WO2023065645A1 WO 2023065645 A1 WO2023065645 A1 WO 2023065645A1 CN 2022091636 W CN2022091636 W CN 2022091636W WO 2023065645 A1 WO2023065645 A1 WO 2023065645A1
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WO
WIPO (PCT)
Prior art keywords
area
concentration
sample
homogenization
dilution
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/CN2022/091636
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English (en)
Chinese (zh)
Inventor
杨柳青
张东锋
吴玥
王超
刘聪
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Autobio Labtec Instruments Zhengzhou Co Ltd
Original Assignee
Autobio Labtec Instruments Zhengzhou Co Ltd
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Publication date
Application filed by Autobio Labtec Instruments Zhengzhou Co Ltd filed Critical Autobio Labtec Instruments Zhengzhou Co Ltd
Priority to EP22882270.6A priority Critical patent/EP4420781A4/fr
Publication of WO2023065645A1 publication Critical patent/WO2023065645A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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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
    • B01L3/50273—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 characterised by the means or forces applied to move the fluids
    • 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
    • B01L3/502769—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 characterised by multiphase flow arrangements
    • B01L3/502784—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 characterised by multiphase flow arrangements specially adapted for droplet or plug flow, e.g. digital microfluidics
    • B01L3/502792—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 characterised by multiphase flow arrangements specially adapted for droplet or plug flow, e.g. digital microfluidics for moving individual droplets on a plate, e.g. by locally altering surface tension
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/06—Fluid handling related problems
    • B01L2200/0605—Metering of fluids
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/06—Fluid handling related problems
    • B01L2200/0694—Creating chemical gradients in a fluid
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/16—Reagents, handling or storing thereof
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0809—Geometry, shape and general structure rectangular shaped
    • B01L2300/0816—Cards, e.g. flat sample carriers usually with flow in two horizontal directions
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/16—Surface properties and coatings
    • B01L2300/161—Control and use of surface tension forces, e.g. hydrophobic, hydrophilic
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/04—Moving fluids with specific forces or mechanical means
    • B01L2400/0403—Moving fluids with specific forces or mechanical means specific forces
    • B01L2400/0415—Moving fluids with specific forces or mechanical means specific forces electrical forces, e.g. electrokinetic
    • B01L2400/0427—Electrowetting

Definitions

  • the invention relates to a microfluidic chip based on electrowetting, in particular to a microfluidic chip and a concentration uniformization method based on electrowetting.
  • Mixed detection of multiple samples has the advantages of saving time, reducing costs, and improving efficiency. It is a commonly used detection method in the fields of gene sequencing, pharmaceutical synthesis, biochemical analysis and diagnosis. Before the mixed sample detection, due to the inconsistent initial concentration of each sample, the concentration of some samples in the same mixed system will be too high after mixing, and the concentration of some samples will be too low. The signal of low concentration samples is easily covered and lost, and the signal of high concentration samples is easy to exceed the detection range, resulting in inaccurate overall detection. Concentration homogenization is a sample mixing method that mixes samples of different concentrations according to a certain ratio to keep the quality or concentration of each sample consistent in the same mixing system. This method can effectively solve the above problems.
  • the concentration homogenization workflow mainly includes three steps: concentration quantification, calculation and comparison, and accurate sample mixing: 1) Concentration quantification: use quantitative detection instruments to detect the initial concentration of different samples; 2) Calculation and comparison: quantify according to the concentration Calculate the volume required for different samples based on the result and the required value of the homogenized concentration; 3) Accurate sample mixing: According to the calculation result, accurately pipette the required volume from each sample, and then mix.
  • Microfluidic systems are widely used in chemical analysis, biomedicine, food hygiene and the environment because of their advantages such as less sample consumption, fast response, high detection efficiency, good heat and mass transfer effects, no cross-contamination, and easy integration with other technical equipment. monitoring and other fields.
  • electrowetting English Electrowetting, abbreviated as EW
  • pipetting technology is a new droplet manipulation method for electrically controlling the surface tension change of the droplet, that is, by changing the voltage between the droplet and the insulating substrate.
  • To change the wettability of the droplet on the substrate that is, to change the contact angle, to promote the deformation and displacement of the droplet.
  • Electrowetting pipetting technology breaks through the pipetting system's dependence on traditional robotic arms, pump valve pipelines and complex flow channels, and can achieve highly flexible and precise control of reagents through digital programming. Therefore, how to use the microfluidic chip to control the concentration of the sample in the microchannel of the microfluidic chip through the electrowetting pipetting technology has not yet been reported.
  • the purpose of the present invention is to provide a microfluidic chip for uniform concentration based on electrowetting. Another purpose of the present invention is to provide a method for using the microfluidic chip to uniformize the concentration, so as to solve the problem that the current manual pipetting is limited by personnel. Operating proficiency, as well as the problems of large pipetting deviation, low accuracy and low efficiency caused by the inherent deviation of the pipette.
  • the present invention takes the following technical solutions:
  • the electrowetting-based concentration uniform microfluidic chip of the present invention includes a microfluidic chip body provided with a microchannel, and the microchannel is used to transport the liquid in it along the microchannel by using electrowetting liquid transfer technology.
  • the microfluidic chip body is also provided with a sample area, a reagent storage area, a dilution area, a concentration quantification area, an accurate sampling area and a homogenization area; the sample area, reagent storage area, dilution area, concentration quantification area, The precise sampling area and the homogenization area communicate through microchannels; where:
  • the sample area is used to store the original sample
  • the reagent storage area is used to store reagents, and the reagents are used to dilute the original sample according to the set gradient;
  • the dilution zone is used to mix the reagent and the original sample entering this zone through the microchannel to prepare a diluted sample;
  • the concentration quantification area is used for concentration determination of the diluted sample entering this area through the microchannel;
  • the total volume and concentration required for homogenization according to the set concentration, and according to the detected or calculated concentration of the original sample and the diluted sample select the appropriate original sample or diluted sample as the mother solution, and calculate the concentration for normalization
  • For the required volume of mother liquor use electrowetting pipetting technology to pipette a set volume of mother liquor to this area to complete concentration homogenization.
  • the microfluidic chip body includes an upper substrate and a lower substrate arranged at an interval between the upper and lower substrates, the microchannel through which the liquid moves is formed between the upper and lower substrates; the structure of the upper substrate from top to bottom is The upper insulating substrate, the common electrode, and the upper hydrophobic layer; the structure of the lower substrate is, from bottom to top, the lower insulating substrate, the driving electrode array, the dielectric layer, and the lower hydrophobic layer.
  • the concentration uniformization method of the concentration uniformization microfluidic chip of the present invention comprises the following steps:
  • Step 1 adding multiple original samples to respective sample areas
  • Step 2 adding the reagent to the reagent storage area
  • Step 3 using electrowetting pipetting technology, moving the reagents to each dilution area according to the set volume, and moving each original sample to the respective dilution area through the microchannel according to the set volume, mixing Prepare diluted samples evenly, and complete the set gradient dilution in sequence;
  • Step 4 using electrowetting pipetting technology, sequentially manipulate the diluted samples in each dilution area, move to the concentration quantification area according to the set volume for electrochemical or fluorescence detection, and directly obtain or calculate the final sample according to the test results.
  • Step 5 according to the total volume and concentration required for homogenization, select the original sample or diluted sample of appropriate concentration as the mother liquor, and calculate the required mother liquor volume;
  • Step 6 Use electrowetting pipetting technology to accurately separate the required volume from each mother liquor through the precise sampling area, and move to the homogenization area for mixing to complete the concentration homogenization.
  • the method of the invention has simple operation and high degree of automation, reduces human operation errors, and the homogenization process can be completed within one hour, which saves time and greatly improves detection efficiency.
  • Electrowetting pipetting technology is used to achieve precise pipetting, and the volume deviation is less than 3%, which solves the problem of inaccurate pipetting results caused by the proficiency of the operator or the inherent deviation of the pipette in traditional pipetting.
  • Fig. 1 is a schematic structural view of the microfluidic chip body of the present invention.
  • Fig. 2 is a schematic diagram of the layout of the sample area, reagent storage area, dilution area, concentration quantification area, precise sampling area, homogenization area and microchannels of the present invention.
  • the electrowetting-based concentration uniform microfluidic chip of the present invention includes a microfluidic chip body provided with a microchannel 1, and the microchannel 1 is used to transport its The droplet 2 inside moves along the microchannel 1.
  • the body of the microfluidic chip includes an upper substrate and a lower substrate arranged at intervals between the upper and lower substrates, and a microchannel 1 through which droplets 2 move is formed between the upper and lower substrates;
  • the structure of the upper substrate is an upper insulating substrate 3, a common The electrode 4, the upper hydrophobic layer 5;
  • the lower substrate structure is the lower insulating substrate 6, the driving electrode array 7, the dielectric layer 8, and the lower hydrophobic layer 9 from bottom to top.
  • the microfluidic chip body is also provided with a sample area, a reagent storage area, a dilution area, a concentration quantification area, an accurate sampling area and a homogenization area; a sample area, a reagent storage area, a dilution area, a concentration quantification area, an accurate sampling area and a homogenization area Areas communicate through microchannel 1; where:
  • the sample area is used to store the original sample; the present embodiment sets four sample areas 10.1, 10.2, 10.3, 10.4;
  • the reagent storage area 11 is used for storing reagents, and the reagents are used to dilute the original sample according to the set gradient;
  • the dilution area is used to mix the reagent and the original sample entering this area through the microchannel 1 to prepare a diluted sample; this embodiment takes two gradient dilutions as an example, and each sample area corresponds to two dilution areas, namely: the first The first dilution zone 12.1 and the second dilution zone 12.2; of course, three gradient dilutions or more gradient dilutions can also be selected according to needs;
  • the concentration quantification area 13 is used to measure the concentration of the diluted sample entering this area through the microchannel 1 by electrochemical or fluorescence detection method;
  • the precise sampling area 14 is used to obtain the original sample or/and the diluted sample according to the set volume
  • Homogenization zone 15 the total volume and concentration required for homogenization according to the set concentration, and according to the detected or calculated concentration of the original sample and the diluted sample, select the appropriate original sample or diluted sample as the mother solution, and calculate the concentration uniformity Use electrowetting pipetting technology to pipette the set volume of mother liquor to this area to complete the concentration homogenization.
  • the concentration uniformization method of the concentration uniform microfluidic chip of the present invention comprises the following steps:
  • Step 1 adding four original samples to the sample areas 10.1, 10.2, 10.3, and 10.4 in advance;
  • Step 2 adding the reagent to the reagent storage area 11 in advance;
  • Step 3 use electrowetting pipetting technology to manipulate the original samples in each sample area 10.1, 10.2, 10.3, and 10.4, move them to the respective first dilution areas 12.1 through the microchannel 1 according to the set volume, and then move them to the respective first dilution areas 12.1 according to the set volume.
  • the reagents are moved to the first dilution area 12.1 with a predetermined volume, and mixed to prepare the first diluted sample; then, the electrowetting pipetting technology is used to control the first diluted sample to move to the second diluted sample through the microchannel 1 according to the set volume.
  • Step 4 using electrowetting pipetting technology, sequentially manipulate the second diluted samples in each second dilution zone 12.2, move to the concentration quantification zone 13 according to the set volume for electrochemical or fluorescence detection, and directly obtain or calculate the concentration of the second diluted sample, and calculate the corresponding concentration of the first diluted sample and the original sample according to the dilution factor of the second diluted sample in each second dilution zone;
  • Step 5 according to the total volume and concentration required for homogenization, select the original sample or diluted sample of appropriate concentration as the mother liquor, and calculate the required mother liquor volume;
  • Step 6 using electrowetting pipetting technology, through the precise sampling area 14, accurately separate the required volume from each mother liquor, and move to the homogenization area 15 for mixing to complete the concentration homogenization.

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  • Chemical & Material Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Dispersion Chemistry (AREA)
  • Analytical Chemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Hematology (AREA)
  • Clinical Laboratory Science (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Automatic Analysis And Handling Materials Therefor (AREA)

Abstract

L'invention concerne une puce microfluidique d'homogénéisation de concentration à base d'électro-mouillage et un procédé d'homogénéisation de concentration ; la puce comprend un corps de puce microfluidique pourvu d'un microcanal ; le microcanal est utilisé pour amener, au moyen d'une technologie de pipetage par électro-mouillage, un liquide à se déplacer le long du microcanal ; le corps de puce microfluidique est pourvu d'une région d'échantillon, d'une région de stockage de réactif, d'une région de dilution, d'une région de quantification de concentration, d'une région d'échantillonnage précise et d'une région d'homogénéisation ; et la région d'échantillon, la région de stockage de réactif, la région de dilution, la région de quantification de concentration, la région d'échantillonnage précise et la région d'homogénéisation communiquent les unes aux autres au moyen du microcanal. Le procédé comprend les étapes suivantes : 1. l'ajout d'une pluralité d'échantillons d'origine aux régions d'échantillon respectives ; 2. l'ajout d'un réactif à la région de stockage de réactif ; 3. l'utilisation d'une technologie de pipetage par électro-mouillage pour déplacer le réactif vers la région de dilution, et le déplacement des échantillons d'origine vers la région de dilution pour achever une dilution de gradient d'ensemble ; 4. la manipulation de l'échantillon dilué pour le déplacer vers la région de quantification de concentration pour la mesure ; 5. la sélection, selon l'homogénéisation, d'un échantillon d'origine en tant que liqueur mère ; et 6. la séparation d'un volume requis au moyen de la région d'échantillonnage précise et le déplacement de celui-ci vers la région d'homogénéisation pour achever l'homogénéisation de la concentration.
PCT/CN2022/091636 2021-10-19 2022-05-09 Puce microfluidique d'homogénéisation de concentration à base d'électro-mouillage et procédé d'homogénéisation de concentration Ceased WO2023065645A1 (fr)

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EP22882270.6A EP4420781A4 (fr) 2021-10-19 2022-05-09 Puce microfluidique d'homogénéisation de concentration à base d'électro-mouillage et procédé d'homogénéisation de concentration

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CN202111217576.X 2021-10-19
CN202111217576.XA CN113842962B (zh) 2021-10-19 2021-10-19 基于电润湿的浓度均一化微流控芯片及浓度均一化方法

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CN113842962B (zh) * 2021-10-19 2023-02-17 安图实验仪器(郑州)有限公司 基于电润湿的浓度均一化微流控芯片及浓度均一化方法
CN115283029B (zh) * 2022-08-01 2025-07-22 郑州安图生物工程股份有限公司 利于消除气泡的微流控芯片

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CN113842962B (zh) 2023-02-17
EP4420781A1 (fr) 2024-08-28
CN113842962A (zh) 2021-12-28

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