CN2821566Y - Single cell algae grain size analysis microflow control chip - Google Patents
Single cell algae grain size analysis microflow control chip Download PDFInfo
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- CN2821566Y CN2821566Y CNU2005200843719U CN200520084371U CN2821566Y CN 2821566 Y CN2821566 Y CN 2821566Y CN U2005200843719 U CNU2005200843719 U CN U2005200843719U CN 200520084371 U CN200520084371 U CN 200520084371U CN 2821566 Y CN2821566 Y CN 2821566Y
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- 241000195493 Cryptophyta Species 0.000 title claims abstract description 10
- 238000004458 analytical method Methods 0.000 title abstract description 15
- 239000007788 liquid Substances 0.000 claims abstract description 23
- 239000002245 particle Substances 0.000 claims description 12
- 239000012530 fluid Substances 0.000 claims description 10
- 239000000758 substrate Substances 0.000 claims description 8
- 239000011521 glass Substances 0.000 claims description 3
- 239000000463 material Substances 0.000 claims description 2
- 210000004027 cell Anatomy 0.000 abstract description 19
- 210000000601 blood cell Anatomy 0.000 abstract description 3
- 210000004748 cultured cell Anatomy 0.000 abstract description 3
- 238000005259 measurement Methods 0.000 abstract description 3
- 239000003792 electrolyte Substances 0.000 abstract 1
- 230000010354 integration Effects 0.000 abstract 1
- 238000001514 detection method Methods 0.000 description 7
- VYZAMTAEIAYCRO-UHFFFAOYSA-N Chromium Chemical compound [Cr] VYZAMTAEIAYCRO-UHFFFAOYSA-N 0.000 description 6
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- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 4
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- 230000004323 axial length Effects 0.000 description 3
- 239000000243 solution Substances 0.000 description 3
- 238000003860 storage Methods 0.000 description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 238000009826 distribution Methods 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 238000000034 method Methods 0.000 description 2
- VLTRZXGMWDSKGL-UHFFFAOYSA-N perchloric acid Chemical compound OCl(=O)(=O)=O VLTRZXGMWDSKGL-UHFFFAOYSA-N 0.000 description 2
- 238000007789 sealing Methods 0.000 description 2
- 238000000926 separation method Methods 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 239000012498 ultrapure water Substances 0.000 description 2
- 239000002699 waste material Substances 0.000 description 2
- RZVAJINKPMORJF-UHFFFAOYSA-N Acetaminophen Chemical compound CC(=O)NC1=CC=C(O)C=C1 RZVAJINKPMORJF-UHFFFAOYSA-N 0.000 description 1
- 238000001712 DNA sequencing Methods 0.000 description 1
- 238000012408 PCR amplification Methods 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- OZECDDHOAMNMQI-UHFFFAOYSA-H cerium(3+);trisulfate Chemical compound [Ce+3].[Ce+3].[O-]S([O-])(=O)=O.[O-]S([O-])(=O)=O.[O-]S([O-])(=O)=O OZECDDHOAMNMQI-UHFFFAOYSA-H 0.000 description 1
- 229910000333 cerium(III) sulfate Inorganic materials 0.000 description 1
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/12—Unicellular algae; Culture media therefor
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
- G01N15/10—Investigating individual particles
- G01N15/14—Optical investigation techniques, e.g. flow cytometry
- G01N15/1429—Signal processing
- G01N15/1433—Signal processing using image recognition
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
- G01N15/10—Investigating individual particles
- G01N15/14—Optical investigation techniques, e.g. flow cytometry
- G01N15/1468—Optical investigation techniques, e.g. flow cytometry with spatial resolution of the texture or inner structure of the particle
- G01N15/147—Optical investigation techniques, e.g. flow cytometry with spatial resolution of the texture or inner structure of the particle the analysis being performed on a sample stream
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
- G01N15/10—Investigating individual particles
- G01N15/14—Optical investigation techniques, e.g. flow cytometry
- G01N15/1484—Optical investigation techniques, e.g. flow cytometry microstructural devices
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Abstract
The utility model discloses a micro-flow control chip for the grain size analysis of single-cell algae, which comprises a base sheet and a cover sheet which are provided with symmetrical structures, wherein a liquid flowing system with a psi type structure comprising a middle sample channel and two side channel sheath flowing channels, and a double-T type detecting channel vertically crossed with the sample channel are arranged on the base sheet and the cover sheet; both sides of the detecting channel are provided with electrodes, and the utility model uses a constant-current detecting system. When a cell passes through a common area for the detecting passage and the sample passage, the cell can replace electrolyte with the same volume as that of the cell to cause resistance between two electrodes to display temporary change, potential difference generates corresponding change to generate pulse signals and then the time and the intensity of the pulse are used for calculating the number and the grain size of the cell. The utility model integrates the Coulter liquid flowing system and the electrodes, incarnates the characteristics of micromation and the integration of a micro-flow control chip system and is suitable for each class of field analysis and real-time measurement. The utility model is used for the fields of single-cell algae, cultured cells, blood corpuscles, etc.
Description
Technical field
The utility model relates to a kind of sreen analysis instrument that utilizes Coulter principle to design and produce, a kind of micro-fluidic chip that is used for the marine unicellular algae sreen analysis of more specifically saying so.
Background technology
Micro-total analysis system (Micro-total analysis system, μ-TAS) proposition of notion has produced significant impact in the analysis science field, and the guiding chemical analysis apparatus is towards the trend development of microminiaturized, integrated and portability.It utilizes micro fabrication to make functional units formation very small chemical systems such as little valve, microchannel, microreactor, microflow sensor, little detecting device on chip.Micro-fluidic chip system has efficiently, low consumption, microminiaturization and integrated characteristics, is suitable for all kinds of on-the site analysiss and The real time measure.Nineteen ninety-five Mathies and Woolley adopt little electrophoresis chip to carry out dna sequencing research first, on the passage of effective separation length 3.5cm, and about 150 bases of order-checking in the 10min, accuracy rate 97%.The microchip gene alaysis system of functions such as integrated cell on micro-fluidic chip such as Ramsey in 1998 is cleared up, pcr amplification and electrophoretic separation.
The principle of Ku Erte particle-size analyzer (Coulter Principle): be suspended in cell or particle in the electrolytic solution, with electrolytic solution during by the aperture pipe, because of having replaced the electrolytic solution of equal volume, in the circuit of continuous current design, cause two resistance between electrode generation transient change inside and outside the aperture pipe, and having produced potential pulse, the size of pulse signal and number of times are directly proportional with the size and the number of particle.Coulter principle belongs to the measurement to the individual particles three-dimensional, therefore, not only can accurately measure cell or particle grain size distribution, more can do the measurement of cell or particle absolute number and concentration, and its institute's particle diameter of surveying is more approaching true.Ku Erte calculating instrument be used widely in fields such as single-cell algae, cultured cell, blood cells (Ray, 2001; Arakawa et al, 1997).
But present Ku Erte calculating instrument costs an arm and a leg, and volume is bigger, is not suitable for on-the site analysis, and the rig-site utilization that appears as Ku Er stunt art of micro-fluidic chip provides may.
Summary of the invention
The purpose of this utility model provides a kind of miniature, portable and be suitable for on-the site analysis marine unicellular algae is counted and the micro-fluidic chip of particle size distribution measuring.
The utility model is based on standard photolithography techniques, is the micro-fluidic chip material with glass, makes up microchannel and little liquid storage tank on chip, makes the sample introduction of sample, detection be integrated on the chip and finish.Micro-fluidic chip comprises the substrate and the cover plate of structural symmetry, is built with to comprise liquid fluid system and the two T-shape sense channels that intersect vertically with sample channel on substrate and cover plate.Liquid fluid system adopts ψ type structure, center-aisle is a sample channel, the subchannel, both sides is called the sheath circulation road, and the effect of sheath circulation road is that the frustule in the sample is arranged with single file, sample channel and sheath circulation road consistent size, the sheath circulation road is 1/4 circular-arc, sample channel is consistent with sheath circulation road end points tangential direction, and like this, sheath stream converges in the same way with sample flow, avoid liquid stream to converge the generation vortex disturbance, guaranteed the stability of liquid fluid system; The end of sample channel and sheath circulation road is built with liquid storage tank respectively, and the other end of sample channel also has a waste liquid pool; On chip, make up the two T-shape sense channels that intersect vertically with sample intake passage, sense channel is designed to semi dumbbell-shape, intersection is carefully lacked (cross sectional dimensions is consistent with sample channel), outside width is big, the resistance of sense channel mainly is the contribution from thin passage, thereby cell just can be obvious by the resistance variations that produces, thereby improve detection sensitivity.Settle electrode in the sense channel both sides, use the continuous current detection system, during the common area of cell by sense channel and sample channel, can replace electrolytic solution with its equal volume, cause that resistance presents temporary change between two electrodes, corresponding change takes place and produces pulse signal in potential difference (PD), calculates cell number and particle diameter by the number of times and the intensity of pulse.
The utility model is integrated with Ku Erte liquid fluid system and electrode, has embodied the microminiaturization of micro-fluidic chip system and integrated characteristics, is suitable for all kinds of on-the site analysiss and The real time measure.Be widely used in fields such as single-cell algae, cultured cell, blood cell.
Description of drawings
Fig. 1 is the structural representation of micro-fluidic chip of the present utility model.
Wherein, 1-sample inlet; 2,3-sheath inflow entrance; 4, the 5-electrode cell; The 6-waste liquid pool; The 7-sample channel; 8,9-sheath circulation road; 10-passage joint; The double-T shaped sense channel of 11-.
Fig. 2 is the structural representation of single-cell algae sreen analysis chip system.
Wherein, 12, the 13-micropump; The 14-constant-current system; 15,16-sheath stream; The 17-sample.
Embodiment
Further specify the utility model below in conjunction with accompanying drawing and by specific embodiment.
Embodiment 1: the structure of micro-fluidic chip
As shown in Figure 1, microfluidic chip structure divides two parts: liquid fluid system and detection system.Liquid fluid system adopts ψ type structure, center-aisle is a sample channel 7, the subchannel, both sides is sheath circulation road 8 and 9, and the effect of sheath circulation road is to make the frustule in the sample be single file arrangement, sample channel 7 and sheath circulation road 8 and 9 consistent size, the sheath circulation road is 1/4 circular-arc, sheath circulation road 8 and 9 consistent with sample channel 7 directions in the tangential direction of joint 10, like this, sheath stream converges in the same way with sample flow, avoid liquid stream to converge the generation vortex disturbance, guaranteed the stability of liquid fluid system; On chip, make up the two T-shape sense channels 11 that intersect vertically with sample channel, sense channel is designed to semi dumbbell-shape, link to each other parts of fine with sample channel 7 and lack, outside width is big, sense channel 11 is 150-200 μ m with the common-use tunnel Design of length of sample channel 7, like this, the resistance of sense channel mainly is the contribution from thin passage, thereby cell just can be obvious by the resistance variations that produces. Electrode cell 4 and 5 places at the sense channel two ends settle electrode, use the continuous current detection system, during the common area of cell by sense channel and sample channel, can replace electrolytic solution with its equal volume, cause that resistance presents temporary change between two electrodes, corresponding change takes place and produces pulse signal in potential difference (PD), calculates cell number and particle diameter by the number of times and the intensity of pulse.
Embodiment 2: the making of micro-fluidic chip
1. the making of glass substrate: the size design of microchannel on the mask film: fluid passage, 30 μ m; Sense channel, 30 μ m, 300 μ m, wide minister 5mm, narrow minister 0.2mm; The long 0.35mm of common-use tunnel.The mask film is placed on the even glue chromium plate of 63mm * 63mm * 1.5mm, ultraviolet exposure 180 seconds (wavelength 365nm) after developing 100 seconds in the developer solution, is dried half an hour down for 100 ℃.At room temperature use chromium film etching liquid (cerous sulfate: perchloric acid: water=50 grams: 15 milliliters: 300 milliliters) corrode the chromium film, rinse oven dry then with high purity water well.By the digit microscope shooting, the channel size that records on the chromium plate is: sample channel, 40 μ m; Sense channel, 40 μ m, 310 μ m.Use 0.5M HF/0.5MNH
4The exposed Pyrex of F etching agent corrosion, speed is about 10 μ m/h, and etching is removed residual light glue-line and chromium film with acetone, chromium film etching liquid after 7 hours more successively, promptly gets substrate.With miniature bench drill punching, drill bit is the diamond bit of 1mm, and the diameter in hole is liquid storage tank diameter 1mm.Microscopically is measured the microchannel size: sample channel, last bottom width 180 μ m, following bottom width 50 μ m, the degree of depth 70 μ m; Sense channel is gone up bottom width 450 μ m, following bottom width 320 μ m, the degree of depth 70 μ m for wide; Common-use tunnel is about 200 μ m
2. the making of cover plate: identical with the substrate method for making, construct identically, but do not punch.
With substrate and cover plate successively at acetone, H
2O-H
2O
2-NH
4OH (5: 1: 1) solution, H
2SO
4: H
2O
2In (4: 1) solution and the high purity water ultrasonic cleaning 5-10 minute, dry up the sealing of in super-clean environment, both being alignd then, bonding under the high temperature with nitrogen.Heating schedule is: rise to 550 ℃ with 40 ℃/min from room temperature, 30 minutes time; Rise to 610 ℃, 30 minutes time with 20 ℃/min from 550 ℃; Rise to 635 ℃, 30 minutes time with 20 ℃/min from 610 ℃; Rise to 650 ℃, 6 hours time with 10 ℃/min from 635 ℃.Naturally cool to room temperature then.Through microscopically observation, passage does not have distortion behind the bonding, and reaches sealing fully.Chip channel xsect ovalize behind the bonding, the narrow minister's axial length of sample channel and sense channel is about 180 μ m, and narrow axial length is about 140 μ m, the about 450 μ m of the wide minister's axial length of sense channel, minor axis is about 140 μ m; Common-use tunnel is about 200 μ m
Embodiment 3: micro-fluidic chip analyzing and testing system
As shown in Figure 2, connect sampling pump 13 in the import 1 of sample channel 7, both sides sheath circulation road 8 and 9 connects sheath stream 15,16 at its inlet 2 and 3 places, settles electrodes to be attempted by on the continuous current meter at sense channel 11 two ends 4,5, constitutes whole micro-fluidic chip analyzing and testing system.Sample and sheath flow liquid are carried by pump 12 and 13, sheath flow liquid is with identical flow rate, it is mobile forward with the form of laminar flow that sheath flow liquid and sample converge the back, by regulating sheath stream and sample flow rate, making cells in sample be single file arranges, and be advanced through detection zone (common area of sense channel and sample channel), replaced isopyknic electrolytic solution at the surveyed area cell, cause that resistance presents temporary change between two electrodes, because detection system is the continuous current design, so the also temporary change of potential difference (PD) produces pulse signal, number and particle diameter that can pair cell according to the number of times of pulse signal and intensity carry out statistical study.
The utility model has made up on the micro-fluidic chip that utilizes the Ku Erte ultimate principle to design and produce realizes required liquid fluid system and the sense channel of Ku Er stunt art, has realized the microminiaturization of Ku Erte sreen analysis system, integrated and simplification; Low cost of manufacture is easy to realize standardization and large-scale production.
Claims (4)
1. particle analyzing chip with microflow control of single-cell algae, it is characterized in that it comprises the substrate and the cover plate of structural symmetry, on substrate and cover plate, be built with the liquid fluid system of the ψ type structure that comprises middle sample passage and two side line sheath circulation roads and the double-T shaped sense channel that intersects vertically with sample channel.
2. micro-fluidic chip according to claim 1 is characterized in that described sheath circulation road is 1/4 circular-arc, and sample channel is consistent with sheath circulation road go side point tangential direction.
3. micro-fluidic chip according to claim 1 is characterized in that described sense channel is designed to semi dumbbell-shape, links to each other parts of fine with sample channel and lacks, and outside width is big.
4. micro-fluidic chip according to claim 1 is characterized in that it is material with glass.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNU2005200843719U CN2821566Y (en) | 2005-06-19 | 2005-06-19 | Single cell algae grain size analysis microflow control chip |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNU2005200843719U CN2821566Y (en) | 2005-06-19 | 2005-06-19 | Single cell algae grain size analysis microflow control chip |
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| CN2821566Y true CN2821566Y (en) | 2006-09-27 |
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| CNU2005200843719U Expired - Fee Related CN2821566Y (en) | 2005-06-19 | 2005-06-19 | Single cell algae grain size analysis microflow control chip |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104140926A (en) * | 2014-07-15 | 2014-11-12 | 大连医科大学附属第二医院 | Device and method for full-automatically sorting circulating tumor cells on micro-fluidic chip |
| CN106190829A (en) * | 2016-07-26 | 2016-12-07 | 西安交通大学 | A kind of for cell high accuracy arrangement and the microflow controlled biochip of detection |
| CN107213930A (en) * | 2017-07-27 | 2017-09-29 | 深圳中科芯海智能科技有限公司 | A kind of micro-fluidic chip and particle analysis method for particle analysis |
| CN108303365A (en) * | 2017-01-13 | 2018-07-20 | 爱科来株式会社 | Flow cell and measurement device |
-
2005
- 2005-06-19 CN CNU2005200843719U patent/CN2821566Y/en not_active Expired - Fee Related
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104140926A (en) * | 2014-07-15 | 2014-11-12 | 大连医科大学附属第二医院 | Device and method for full-automatically sorting circulating tumor cells on micro-fluidic chip |
| CN106190829A (en) * | 2016-07-26 | 2016-12-07 | 西安交通大学 | A kind of for cell high accuracy arrangement and the microflow controlled biochip of detection |
| CN106190829B (en) * | 2016-07-26 | 2018-07-03 | 西安交通大学 | A kind of microflow controlled biochip for arranging and detecting for cell high-precision |
| CN108303365A (en) * | 2017-01-13 | 2018-07-20 | 爱科来株式会社 | Flow cell and measurement device |
| CN108303365B (en) * | 2017-01-13 | 2022-06-24 | 爱科来株式会社 | Flow cell and assay device |
| CN107213930A (en) * | 2017-07-27 | 2017-09-29 | 深圳中科芯海智能科技有限公司 | A kind of micro-fluidic chip and particle analysis method for particle analysis |
| CN107213930B (en) * | 2017-07-27 | 2022-12-20 | 深圳中科芯海智能科技有限公司 | Microfluidic chip for particle analysis and particle analysis method |
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