CN101561448B - Negative-pressure pinched injection method of micro-fluidic chip based on integrated minipump valve and special chip thereof - Google Patents
Negative-pressure pinched injection method of micro-fluidic chip based on integrated minipump valve and special chip thereof Download PDFInfo
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- CN101561448B CN101561448B CN2008100110858A CN200810011085A CN101561448B CN 101561448 B CN101561448 B CN 101561448B CN 2008100110858 A CN2008100110858 A CN 2008100110858A CN 200810011085 A CN200810011085 A CN 200810011085A CN 101561448 B CN101561448 B CN 101561448B
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Abstract
The invention provides a negative-pressure pinched injection method of a micro-fluidic chip based on an integrated minipump valve and a special chip thereof. The injection method comprises the following steps: adding a buffer solution reagent to a buffer solution storage bath, a buffer waste solution bath and a sample waste solution bath; putting a sample to be detected into a sample bath; carrying out the following steps by pump valves: combining four valves into two groups of pumps to run by programmed control, generating negative pressure by the elastic deformation of a PDMS film, enabling liquid in the sample bath, the buffer solution storage bath and the buffer waste solution bath to flow to the sample waste solution bath under the action of the negative pressure so as to form a pinched sample section belt; and exerting a voltage between the buffer solution storage bath and the buffer waste solution bath for electrophoretic separation and detection. The invention has the advantages of short injection time, no sample discrimination effect, good reproducibility, easy operation, integration, and the like.
Description
Technical field
The present invention relates to chemistry and physics, be specifically related to the chip electrophoresis technology, the pump valve integrated technology on the chip provides the negative-pressure pinched injection method of micro-fluidic chip based and the special chip of integrated Micropump valve especially.
Background technology
Along with differential compartment analysis technology rapid development, microflow control technique has obtained people's vast concern as an important techniques in the more than ten years recently, and its principal feature is that the flexible combination and the scale of various functional units is integrated.And the sample feeding technology has also obtained the broad research of researchers as most important technological means in the micro-fluidic chip compartment analysis.
The micro-fluidic chip technology is the research focus of current instrumental analysis; This technology mainly is the basis with analytical chemistry and biological chemistry; Be applied to various fields such as chemical analysis, drug screening, immunoassay, medical diagnosis on disease, cell analysis; And sampling technique has received a large amount of concerns of this area researchist as a guardian technique in the micro-fluidic chip platform.
In existing research, be divided into simple sampling system again according to the difference of sample injection method, T passage sample introduction, double T passage sample introduction, extraining sampling etc., and be divided into electrokinetic injection and pressure sample introduction etc. according to the different sampling techniques of driving force.
1, adopting cross passage sample introduction is method the most commonly used in the chip capillary cataphoresis, can realize the sample introduction operation through the switching of voltage between specimen access and split tunnel.Its principle of operation is at first between sample cell and sample waste liquid pool, to apply voltage, and sample is full of sample intake passage under the effect of EOF, and making alive is carried out separation detection in split tunnel then.
2, in order to increase sample size; Can change the cruciform passage into the double T passage; But these two kinds of methods all have the effect of electrical discrimination to sample, must need time enough to wait until that the minimum component of migration rate also arrives right-angled intersection injection port place and could represent real sample to form.
3, electronic extraining sampling is to make sample waste liquid pool ground connection; Its excess-three liquid pool applies high voltage respectively then, and flow can flow to the sample waste liquid pool from three liquid pools under the effect of electric field force, thereby makes the sample area band form folder stream; In split tunnel, apply voltage then; Realize the separation and the detection of sample, this method has the separation efficiency height, advantages such as n.s. leakage.
Above-mentioned method all exists the effect of electrical discrimination of sample; Therefore use pressure to replace the method for electric power sample introduction more and more to receive researchist's concern and attention, the pressure sample injection method comprises methods such as static pressure sample introduction, gravity sample introduction, external pump valve realization pressure sample introduction.When driving force is pressure, also can realize the extraining sampling under the pressure effect.
People expect to obtain a kind of be easy to integrated, sample introduction and do not have sample the effect of electrical discrimination, sample injection method and corresponding apparatus thereof on the micro-fluidic chip of characteristics such as reappearance height fast.
Summary of the invention
The purpose of this invention is to provide the negative-pressure pinched injection method of micro-fluidic chip based and the special chip of integrated Micropump valve, this chip has the sample introduction process and does not have sample the effect of electrical discrimination, and reappearance is high, and the working cell has integrated advantage.
The invention provides the negative pressure extraining sampling method of the micro-fluidic chip of integrated Micropump valve, concrete steps are:
Required damping fluid reagent is put into damping fluid liquid storage tank, damping fluid waste liquid pool and sample waste liquid pool, and the sample that needs to detect is put into sample cell;
Carry out the sample introduction step through pump valve: two groups of pumps being made up of 4 valves move through programmed control; Elastic deformation by the PDMS film produces negative pressure; Under action of negative pressure; Flow direction sample waste liquid pool in sample cell, damping fluid liquid storage tank, the damping fluid waste liquid pool forms the sample area band that folder flows;
Between damping fluid liquid storage tank and damping fluid waste liquid pool, apply voltage, carry out electrophoretic separation and detection.
The negative pressure extraining sampling method of the micro-fluidic chip based on integrated Micropump valve provided by the invention, employed damping fluid normally is fit to the required damping fluid of separation condition with a kind of ability in the different separative element in the said micro-fluidic chip; Electrophoresis process damping fluids more commonly used are as operation solution, for example phosphate buffer, borate buffer solution and acetate buffer solution etc.
The negative pressure extraining sampling method of the micro-fluidic chip based on integrated Micropump valve provided by the invention; Said at valve 5; 6 open when forming the sample area band that presss from both sides stream, between damping fluid liquid storage tank and damping fluid waste liquid pool, apply, and field strength range is 300～800 volts/centimetre.
The negative pressure extraining sampling method of the micro-fluidic chip based on integrated Micropump valve provided by the invention, when adopting the laser induced fluorescence detector of point focusing formula to detect, the concentration of the sample compound that needs to detect is 10
-6～10
-7Mol.
The present invention also provides a kind of micro-fluidic chip that is used for the integrated Micropump valve of negative pressure extraining sampling method; This micro-fluidic chip is the chip of glass-PDMS-glass hybridization; Formed by upper strata, lower floor and middle layer three part sealing-ins, two-layer up and down is glass material, and microfluid passage is contained on the upper strata; The middle layer is the PDMS film of 254um, and gas channels is contained in lower floor.
The micro-fluidic chip of the integrated Micropump valve that is used for negative pressure extraining sampling method that the present invention also provides; This micro-fluidic chip is externally connected to the solenoid valve of program controlled; Under the control of the program of finishing in advance, the pressure in can switching gas circuit, deformation takes place in PDMS under the effect of pressure; Realize the open and close function of valve, the valve of three this structures realizes that under certain sequence of operation the liquid transportation is the function of pump.
The micro-fluidic chip that is used for the integrated Micropump valve of negative pressure extraining sampling method provided by the invention, liquid road, said upper strata layer is made up of sample cell, damping fluid liquid storage tank, damping fluid waste liquid pool, sample waste liquid pool, little valve, split tunnel; The number of little valve is 3～10 in the chip.
The micro-fluidic chip that is used for the integrated Micropump valve of negative pressure extraining sampling method provided by the invention, the degree of depth of said split tunnel or width are 10-100um.
The micro-fluidic chip that is used for the integrated Micropump valve of negative pressure extraining sampling method provided by the invention, said lower floor gas circuit layer is made up of valve seat, the gentle way outlet of gas channels.
The micro-fluidic chip that is used for the integrated Micropump valve of negative pressure extraining sampling method provided by the invention, the width of said gas channels or the degree of depth are 50-250um, and vacuum tightness is 40-80KPa, and pressure is 10-100KPa.
The micro-fluidic chip that is used for the integrated Micropump valve of negative pressure extraining sampling method provided by the invention, reversible sealing-in PDMS film in the middle of said upper strata microfluid passage chip and the lower floor's gas channels chip, test earlier guarantees that whole valves can operate as normal before experiment.
Design to the invention property the micro-fluidic chip of the negative pressure extraining sampling that is used for little integrated pump valve, can realize the efficient sample introduction fast that n.s. is discriminated against.Whole process was accomplished in one minute, and sample consumption is at micro updating.The present invention compares with classic method, has the short and n.s. of sample injection time and discriminates against effect, favorable reproducibility height, easy operating, advantage such as integrated.
The present invention can accomplish the compartment analysis of sample at short notice on more than one square centimeters plastics, glass and PDMS hybridization hybrid chip.In terms of existing technologies, the present invention have the short and without discrimination effect of sample injection time, integrated, be easy to characteristics such as control operation and amount of samples are few.
Description of drawings
The negative pressure extraining sampling microfluidic chip structure figure of Fig. 1 integrated Micropump valve of the present invention;
Liquid road, the upper strata chip structure figure of this chip of Fig. 2, wherein: 1 sample cell, 2 damping fluid liquid storage tanks, 3 damping fluid waste liquid pools, 4 sample waste liquid pools, 5,6,7,8 little valves, 9 split tunnels;
The gas circuit chip structure figure of lower floor of this chip of Fig. 3, wherein: 10,11,12,13 valve seats, 14,15,16,17 gas channels, the outlet of 18,19,20,21 gas circuits;
Fig. 4 is sample real-time fluorescence spectrogram with the uranin;
This chip of Fig. 5 is to 25 spectrograms of uranin sample continuous sample introduction;
Separating wire propylhomoserin and glycocoll spectrogram on this chip of Fig. 6.
Embodiment
Following embodiment will further explain the present invention, but therefore not limit the present invention.
Embodiment 1: the making of micro-fluidic chip
Micro-fluidic chip (like Fig. 1) is that the liquid road layer that is etched with microfluid passage on the upper strata, gas circuit layer and the middle reversible sealing-in of PDMS film that accompanies 254um that lower floor is etched with gas channels form.The liquid paths degree of depth and width are respectively 20um and 80um; The gas channels degree of depth and width are respectively 80um and 200um, and be firm for guaranteeing sealing-in, needs to keep chip clean; Need to aim at sealing-in in microscopically, and guarantee the operate as normal of valve at the ultra-clean chamber of 10000 cleanliness factors.The place connects peripheral gas circuit at the gas channels mouth, realizes the switching of positive/negative pressure in the gas channels through the switch of setting program control electromagnetic valve in advance, and three valves can be realized the function of pump under the motion of a definite sequence.Embodiment 2: the negative pressure sample introduction research that with the uranin is the integrated Micropump valve of chip of object
In the sample liquid pool of embodiment 1 described micro-fluidic chip, add Fluress 1 * 10
-6M, all the other liquid pools add the borax buffer solution of PH=9.2, control the motion of little valve; Produce the pressure of negative sense, under the effect of pressure, the liquid in sample cell, buffer pool, the sample waste liquid pool is inhaled in the valve; And separation detection behind the generation folder stream sample area band, as shown in Figure 4.After obtaining Stable Manifold, apply 500 volts of/centimetre voltages at split tunnel, make its separation detection.This method can realize the analysis of rapid Continuous sample feeding, accomplishes an analysis operation in 25 seconds, and the result is as shown in Figure 5.
Embodiment 3:
To amino acid whose analyzing and testing.Present embodiment implementation process and embodiment 2 basically identicals, difference is that institute's analytic target is the amino acid sample.The amino acid that present embodiment adopted is serine (Serine) and glycocoll (Glycine), at first carries out mark with FITC, and the mol ratio of amino acid and FITC is 10: 1; Amino acid is excessive; Analyze with said method, but two seed amino acid baseline separation are as shown in Figure 6.Advantages such as The whole analytical process has been eliminated the shortcoming that the sample electricity is discriminated against sample introduction, and is automatical and efficient.
Claims (5)
1. negative pressure extraining sampling method based on the micro-fluidic chip of integrated Micropump valve, it is characterized in that: concrete steps are:
Required damping fluid reagent is put into damping fluid liquid storage tank (2), damping fluid waste liquid pool (3) and sample waste liquid pool (4), and the sample that needs to detect is put into sample cell (1);
Carry out the sample introduction step through pump valve: two groups of pumps being made up of 4 valves move through programmed control; Elastic deformation by the PDMS film produces negative pressure; Under action of negative pressure; Flow direction sample waste liquid pool in sample cell (1), damping fluid liquid storage tank (2), the damping fluid waste liquid pool (3) forms the sample area band that folder flows;
Between damping fluid liquid storage tank (2) and damping fluid waste liquid pool (3), apply voltage, field strength range is 300～800 volts/centimetre, carries out electrophoretic separation and detection.
2. according to the negative pressure extraining sampling method of the said micro-fluidic chip based on integrated Micropump valve of claims 1, it is characterized in that: when adopting the laser induced fluorescence detector of point focusing formula to detect, the concentration of the sample compound that needs to detect does
10
-6～10
-7Mol.
3. micro-fluidic chip that is used for the integrated Micropump valve of the said negative pressure extraining sampling of claim 1 method; It is characterized in that: this micro-fluidic chip is the chip of glass-PDMS-glass hybridization; Formed by upper strata, lower floor and middle layer three part sealing-ins, two-layer up and down is glass material, and microfluid passage is contained on the upper strata; The middle layer is the PDMS film of 254um, and gas channels is contained in lower floor;
Liquid road, upper strata layer is made up of sample cell (1), damping fluid liquid storage tank (2), damping fluid waste liquid pool (3), sample waste liquid pool (4), little valve (5,6,7,8), split tunnel (9);
Lower floor's gas circuit layer is made up of valve seat (10,11,12,13), the gentle way outlet of gas channels (14,15,16,17) (18,19,20,21);
Reversible sealing-in PDMS film in the middle of upper strata microfluid passage chip and the lower floor's gas channels chip.
4. according to the said micro-fluidic chip that is used for the integrated Micropump valve of negative pressure extraining sampling method of claim 3, it is characterized in that: the degree of depth of said split tunnel (9) or width are 10-100um.
5. according to the said micro-fluidic chip that is used for the integrated Micropump valve of negative pressure extraining sampling method of claim 3, it is characterized in that: said gas channels (14,15; 16; 17) the width or the degree of depth are 50-250um, and vacuum tightness is 40-80KPa, and pressure is 10-100KPa.
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| CN102161967A (en) * | 2010-02-24 | 2011-08-24 | 中国科学院大连化学物理研究所 | Microfluidic-based nucleic acid hybridization reaction platform and hybridization analysis method |
| CN101773861B (en) * | 2010-04-02 | 2011-09-14 | 华中科技大学 | Microfluidic sample feeding method, device and application thereof |
| CN102262162B (en) * | 2010-05-26 | 2013-05-29 | 中国科学院大连化学物理研究所 | Microfluidic chip system for studying mechanical behaviors of cells |
| CN103146574B (en) * | 2011-12-07 | 2015-09-30 | 国家纳米科学中心 | A kind of high-throughput micro-fluidic biological mechanics long-time stimulus system and application thereof |
| CN102671729B (en) * | 2012-05-07 | 2014-04-16 | 博奥生物有限公司 | Micro-fluidic chip for multi-index biochemical detection |
| CN104415801B (en) * | 2013-08-28 | 2016-03-09 | 中国科学院青岛生物能源与过程研究所 | A kind of portable microlayer model generator based on self energizing vacuum Micropump |
| CN105707010A (en) * | 2014-12-05 | 2016-06-29 | 中国科学院大连化学物理研究所 | Micro-valve-based dual-detection microfluidic chip and diabetic model construction method |
| CN105734045B (en) * | 2014-12-11 | 2019-02-01 | 中国科学院大连化学物理研究所 | A method of the quick multi-pass amount based on micro-fluidic chip extracts blood sample DNA |
| CN105733923B (en) * | 2014-12-11 | 2018-06-19 | 中国科学院大连化学物理研究所 | A kind of micro-fluidic chip and the nucleic acid extraction purification process using micro-fluidic chip |
| CN104675808B (en) * | 2015-03-09 | 2017-08-04 | 哈尔滨工程大学 | A kind of optical fiber microfluid drive and driving method |
| CN106959332B (en) * | 2017-02-28 | 2019-03-29 | 中国科学院合肥物质科学研究院 | The automatic fine sampling device and its control method of electrophoresis detection instrument |
| CN109012768B (en) * | 2017-06-09 | 2021-11-19 | 国家纳米科学中心 | Microfluidic liquid one-way flow control structure, chip and method |
| CN110423686A (en) * | 2019-07-01 | 2019-11-08 | 上海市第十人民医院 | Cell excretion body optimization culture system and cell excretion body optimization culture method |
| CN113373036A (en) * | 2021-05-31 | 2021-09-10 | 哈尔滨工业大学 | Visual micro-fluidic chip for multi-virus nucleic acid detection and detection method thereof |
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| CN1740779A (en) * | 2005-09-22 | 2006-03-01 | 复旦大学 | Analyzing system and analysing method based on integrated micro-flow control chip |
| CN2898830Y (en) * | 2006-03-14 | 2007-05-09 | 浙江大学 | Microcurrent controlled chip negative-pressure separator with sampling function by miniature vacuum pump |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1740779A (en) * | 2005-09-22 | 2006-03-01 | 复旦大学 | Analyzing system and analysing method based on integrated micro-flow control chip |
| CN2898830Y (en) * | 2006-03-14 | 2007-05-09 | 浙江大学 | Microcurrent controlled chip negative-pressure separator with sampling function by miniature vacuum pump |
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