CN1908647A - Ngatively pressurized sampling three-dimensional chip capillary array electrophoresis system - Google Patents
Ngatively pressurized sampling three-dimensional chip capillary array electrophoresis system Download PDFInfo
- Publication number
- CN1908647A CN1908647A CN 200610053141 CN200610053141A CN1908647A CN 1908647 A CN1908647 A CN 1908647A CN 200610053141 CN200610053141 CN 200610053141 CN 200610053141 A CN200610053141 A CN 200610053141A CN 1908647 A CN1908647 A CN 1908647A
- Authority
- CN
- China
- Prior art keywords
- sample
- public
- swimming lane
- waste liquid
- liquid pool
- 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.)
- Granted
Links
- 238000001962 electrophoresis Methods 0.000 title claims abstract description 46
- 238000005070 sampling Methods 0.000 title claims description 35
- 238000000926 separation method Methods 0.000 claims abstract description 33
- 238000001917 fluorescence detection Methods 0.000 claims abstract description 7
- 239000000523 sample Substances 0.000 claims description 116
- 239000007788 liquid Substances 0.000 claims description 67
- 239000002699 waste material Substances 0.000 claims description 64
- 239000000758 substrate Substances 0.000 claims description 61
- 230000009182 swimming Effects 0.000 claims description 60
- 239000012530 fluid Substances 0.000 claims description 53
- 238000013016 damping Methods 0.000 claims description 31
- 238000005251 capillar electrophoresis Methods 0.000 claims description 27
- 239000000872 buffer Substances 0.000 claims description 18
- 238000003860 storage Methods 0.000 claims description 17
- 239000011521 glass Substances 0.000 claims description 14
- 238000012545 processing Methods 0.000 claims description 13
- 239000012488 sample solution Substances 0.000 claims description 13
- 230000000694 effects Effects 0.000 claims description 10
- 238000000034 method Methods 0.000 claims description 9
- 238000005370 electroosmosis Methods 0.000 claims description 8
- 238000004458 analytical method Methods 0.000 claims description 7
- 230000005684 electric field Effects 0.000 claims description 7
- BASFCYQUMIYNBI-UHFFFAOYSA-N platinum Chemical compound [Pt] BASFCYQUMIYNBI-UHFFFAOYSA-N 0.000 claims description 6
- 230000008859 change Effects 0.000 claims description 5
- 239000007853 buffer solution Substances 0.000 claims description 4
- 238000005520 cutting process Methods 0.000 claims description 3
- 229910052697 platinum Inorganic materials 0.000 claims description 3
- 230000008569 process Effects 0.000 claims description 2
- 230000003287 optical effect Effects 0.000 description 7
- GNBHRKFJIUUOQI-UHFFFAOYSA-N fluorescein Chemical compound O1C(=O)C2=CC=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 GNBHRKFJIUUOQI-UHFFFAOYSA-N 0.000 description 5
- 238000002347 injection Methods 0.000 description 4
- 239000007924 injection Substances 0.000 description 4
- 239000000203 mixture Substances 0.000 description 4
- VYZAMTAEIAYCRO-UHFFFAOYSA-N Chromium Chemical compound [Cr] VYZAMTAEIAYCRO-UHFFFAOYSA-N 0.000 description 3
- 239000004568 cement Substances 0.000 description 3
- 229910052804 chromium Inorganic materials 0.000 description 3
- 239000011651 chromium Substances 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- 238000005530 etching Methods 0.000 description 3
- 238000001499 laser induced fluorescence spectroscopy Methods 0.000 description 3
- 239000011259 mixed solution Substances 0.000 description 3
- TUFFYSFVSYUHPA-UHFFFAOYSA-M rhodamine 123 Chemical compound [Cl-].COC(=O)C1=CC=CC=C1C1=C(C=CC(N)=C2)C2=[O+]C2=C1C=CC(N)=C2 TUFFYSFVSYUHPA-UHFFFAOYSA-M 0.000 description 3
- 238000003556 assay Methods 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 229920000642 polymer Polymers 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 239000010453 quartz Substances 0.000 description 2
- PYWVYCXTNDRMGF-UHFFFAOYSA-N rhodamine B Chemical compound [Cl-].C=12C=CC(=[N+](CC)CC)C=C2OC2=CC(N(CC)CC)=CC=C2C=1C1=CC=CC=C1C(O)=O PYWVYCXTNDRMGF-UHFFFAOYSA-N 0.000 description 2
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N silicon dioxide Inorganic materials O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- -1 Argon ion Chemical class 0.000 description 1
- 238000001712 DNA sequencing Methods 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- XKRFYHLGVUSROY-UHFFFAOYSA-N argon Substances [Ar] XKRFYHLGVUSROY-UHFFFAOYSA-N 0.000 description 1
- 229910052786 argon Inorganic materials 0.000 description 1
- 238000003491 array Methods 0.000 description 1
- 229910021538 borax Inorganic materials 0.000 description 1
- 238000011088 calibration curve Methods 0.000 description 1
- 238000003759 clinical diagnosis Methods 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 230000008878 coupling Effects 0.000 description 1
- 238000010168 coupling process Methods 0.000 description 1
- 238000005859 coupling reaction Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 239000010432 diamond Substances 0.000 description 1
- 229910003460 diamond Inorganic materials 0.000 description 1
- 230000003828 downregulation Effects 0.000 description 1
- 238000007877 drug screening Methods 0.000 description 1
- 229920006335 epoxy glue Polymers 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 230000002349 favourable effect Effects 0.000 description 1
- 238000013467 fragmentation Methods 0.000 description 1
- 238000006062 fragmentation reaction Methods 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 238000013508 migration Methods 0.000 description 1
- 230000005012 migration Effects 0.000 description 1
- 238000001259 photo etching Methods 0.000 description 1
- 238000006552 photochemical reaction Methods 0.000 description 1
- 229920001184 polypeptide Polymers 0.000 description 1
- 102000004196 processed proteins & peptides Human genes 0.000 description 1
- 108090000765 processed proteins & peptides Proteins 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 238000012163 sequencing technique Methods 0.000 description 1
- 239000004328 sodium tetraborate Substances 0.000 description 1
- 235000010339 sodium tetraborate Nutrition 0.000 description 1
- 239000000243 solution Substances 0.000 description 1
- 238000011144 upstream manufacturing Methods 0.000 description 1
Images
Landscapes
- Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)
Abstract
The electrophoresis system comprises: a 3Dcapillary array micro-chip with multiple separation units, a negative feeding and separation device with micro vacuum pump as the negative pressure source, a high-voltage dc power, and a laser fluorescence detection system. This invention reduces cost greatly, and has well repeatability.
Description
Technical field
The present invention relates to the microfluid control chip capillary electrophoresis analysing technology, particularly relate to the making and the sampling technique of microchip capillary array electrophoresis.
Background technology
The micro flow control chip capillary electrophoresis technology has been used for dna sequencing, the separation of dna fragmentation and evaluation, the analysis of the separation determination of amino acid, polypeptide, protein and unicellular interior component etc.But, have only a chip capillary cataphoresis that separates swimming lane can not realize multiple parallel mensuration, be difficult to satisfy needs such as batch samples compartment analysises such as gene sequencing, proteomics research, clinical diagnosis, drug screenings.Development chip capillary array electrophoresis analytic system can be measured by multiple parallel, increases substantially work efficiency, reduces analysis cost.
Mainly adopt electrokinetic injection mode sample introduction at present in chip capillary array electrophoresis, each chip capillary cataphoresis separative element all need be equipped with respective electrode, and it is many more to separate swimming lane, the also proportional increase of electrode number, and corresponding power supply and control system are also complicated more.And owing to " discrimination effect " arranged during electrokinetic injection, promptly negative ions migration velocity in electric field is inconsistent, causes the composition of the composition of sample plug and sample solution inconsistent.Can reduce difference on sample plug and the sample solution composition by prolonging sample injection time, but weaken the characteristics of micro-fluidic chip express-analysis.Simultaneously, the variation of chip capillary surface nature can cause the change of electroosmotic flow size, and the precision of sample introduction is reduced greatly; When the capillary surface change of properties was serious, the electroosmotic flow direction also can change, and made sample can't enter sample introduction and separated swimming lane.
In order to address the above problem, in chip capillary array electrophoresis, adopt the optical gate sample introduction that report is also arranged.The capillary array electrophoresis chip is fixed on the platform moves back and forth.The laser beam that is produced by high-power Argon ion laser is divided into two different bundles of energy by chopper, and the gate light beam that wherein energy is big is focused on the passage upstream near the sample cell place, and the detection light beam that energy is little is focused on the passage downstream near the buffer pool place.Not during sample introduction, fluorescent chemicals in the sample is under the effect of electroosmotic flow, move downstream along linear path from sample cell, when the spot of gate light beam, under high-octane laser action, resolve into fluorescigenic hardly compound (photobleaching) because of photochemical reaction takes place, these non-fluorescent chemicalses have formed stable baseline when flowing through the detection light beam in passage downstream with damping fluid.When the needs sample introduction, block the gate light beam with an optical gate, it no longer is radiated on the passage, at this moment, the fluorescent chemicals in the sample with in its original existence form admission passage, the downstream, and then obtain separating and detect.The size of sample size is determined that by the time that optical gate blocks the gate light beam time is longer, and sample size is bigger, and the signal that records is healed strong, and post is imitated then lower.Because the apparatus expensive of optical gate sample introduction, complex structure, and also sample must have fluorescence, and its application has been subjected to certain limitation.
Number of patent application: 200510050547.4 disclose a kind of Ngatively pressurized sampling and the device that separates, and by micro-fluidic chip, ram pump, T-valve, interface, high-voltage power supply is formed.Number of patent application: 200610048906.6 disclose a kind of micro flow control chip negative pressure sampling and the device that separates with minipump, are made up of micro-fluidic chip, minipump, Dewar bottle, electric contact vacuum meter, three-way solenoid valve, interface and high-voltage power supply.Number of patent application: 200610048907.0 and 200510050458.9 disclose micro flow control chip capillary electrophoresis negative pressure sampling method, it is fast to have sample introduction speed, handling safety, advantages such as equipment is simple, but can only carry out an electrophoretic analysis that separates swimming lane, can not satisfy the needs that chip capillary array electrophoresis is analyzed.
Summary of the invention
The purpose of this invention is to provide a kind of simple and reliable micro flow control chip capillary array electrophoresis system.Overcome present micro flow control chip capillary electrophoresis sample introduction apparatus expensive, the shortcoming that complex structure and sample injection time are long.
Ngatively pressurized sampling three-dimensional chip capillary array electrophoresis system provided by the invention, comprise the three-dimensional capillary array electrophoresis microchip that contains a plurality of capillary electrophoresis separation unit, minipump is as the Ngatively pressurized sampling of negative pressure source and the device that separates, high-voltage DC power supply and laser fluorescence detection system are formed, described three-dimensional capillary array electrophoresis microchip is made by three layers of substrate, and public buffer pool B is arranged on the substrate of upper strata
T, public damping fluid waste liquid pool BW
T, each sample solution pool S
1-S
nWith public sample waste liquid pool SW
TThe channel network that a plurality of capillary electrophoresis separation unit is arranged on the substrate of middle one deck, each separative element separates swimming lane by one and a sample intake passage is formed, and the entrance point that separates swimming lane is by a public damping fluid channel B
BusBe linked together, and pass through at the aperture of substrate corresponding position, upper strata and public buffer pool B
TBe associated in together, the endpiece of separation swimming lane passes through aperture and the damping fluid waste liquid pool BW in the substrate corresponding position on upper strata
TBe connected, the entrance point of every sample intake passage also with each sample solution pool S of substrate corresponding position on upper strata
1-S
nBe connected; A public sample waste fluid channel is arranged on bottom substrate, on-chip aperture links together with the public sample waste fluid channel that is positioned on the bottom substrate sample intake passage endpiece in each separative element in the middle level by processing, and the relevant position aperture by upper strata and middle level glass substrate, make public sample waste fluid channel that is positioned at bottom substrate and the public sample waste liquid pool SW that places the upper strata substrate
TCommunicate; Described Ngatively pressurized sampling device is by minipump, Dewar bottle, electric contact vacuum meter, three-way solenoid valve, timer, and interface is formed, by interface and public sample waste liquid pool SW
TLink to each other; The positive and negative electrode of described high-voltage DC power supply and public buffer pool B
TWith damping fluid waste liquid pool BW
TIn electrophoresis buffer solution be connected; Described laser fluorescence detection system is made up of charge-coupled pick-up unit, color filter, laser instrument and lens.
Electrophoresis system of the present invention comprises a three-dimensional capillary array electrophoresis microchip that contains a plurality of capillary electrophoresis separation unit, one with minipump as the Ngatively pressurized sampling of negative pressure source and the device that separates, a high-voltage DC power supply and a laser fluorescence detection system.Only just can finish the assay determination of array chip capillary electrophoresis analysis system multiple parallel with a micro flow control chip negative pressure sampling and the device that separates and high-voltage DC power supply, greatly reduce the cost of manufacture of array chip capillary electrophoresis system, and easy to operate, safety, sample introduction speed are fast, favorable reproducibility.
The present invention uses a micro flow control chip negative pressure sampling device, when three-way solenoid valve b end and the connection of c end, at public sample waste liquid pool SW
TThe middle negative pressure that forms, sample solution on the micro-fluidic chip in other liquid storage tanks and damping fluid are passed through to process in the on-chip public sample waste fluid channel of lower floor to public sample waste liquid pool SW under atmospheric effect
TFlow, in 0.5 second, form stable sample plug at a plurality of sample intake passages with the infall that separates swimming lane simultaneously.
The present invention is communicated with three-way solenoid valve to switch to the b end and hold with a by b end and c end automatically 0.5 second the time by the timer in the Ngatively pressurized sampling device and is communicated with, and makes public sample waste liquid pool SW
TCommunicate with atmosphere, the pressure differential between it and other liquid pool disappears immediately simultaneously, forms sample plug at a plurality of sample intake passages and the infall that separates swimming lane and is added in the electroosmotic flow carry over score that electric field produced on the separation swimming lane from swimming lane thereby make, and begins electrophoretic separation.
The present invention is in public damping fluid channel B
BusIn placed the platinum filament of 5 microns of diameters, and by at the aperture of substrate corresponding position, upper strata and public buffer pool B
TBe associated in together; The endpiece of separation swimming lane passes through aperture and the damping fluid waste liquid pool BW in the substrate corresponding position on upper strata
TBe connected, with the positive and negative electrode of an adjustable high-voltage DC power supply and public buffer pool B
TWith damping fluid waste liquid pool BW
TIn electrophoresis buffer solution be connected, three-way solenoid valve b end-grain cutting change to a end after, a plurality of sample intake passages and the infall that separates swimming lane form sample plug and are added in the identical electroosmotic flow carry over score that electric field produced that separates on the swimming lane from swimming lane, carry out replicate determination in a plurality of separation swimming lanes simultaneously.
The present invention makes in the array chip capillary electrophoresis chip a large amount of capillary electrophoresis separation swimming lanes and public sample waste fluid channel processing on two different planes. and use micro flow control chip negative pressure sampling method (number of patent application: 200610048907.0 and 200510050458.9), sample introduction and replicate determination when just can finish a plurality of sample of array chip Capillary Electrophoresis.
Advantage of the present invention and effect are as follows:
1. when the present invention selects for use the Ngatively pressurized sampling apparatus and method to realize in the capillary array electrophoresis microchip a large amount of capillary electrophoresis separation unit sample introduction with separate, have that sampling device is simple in structure, with low cost, easy to operate, negative regulation is controlled, sample introduction speed is fast, do not have " discrimination effect ", just can obtain in 0.5 second with the sample liquid storage tank in the identical sample plug of solution composition.
2. the present invention is by processing Capillary Electrophoresis function channel array and public sample waste fluid channel respectively on middle level and lower floor's substrate, again by last, in two substrate relevant positions methods of holing, made three-dimensional capillary array electrophoresis microchip, the sample waste liquid pool links together in each separative element thereby make, and only just can finish the assay determination of quick sample introduction of array chip capillary electrophoresis analysis system and multiple parallel with a micro flow control chip negative pressure sampling and the device that separates and high-voltage DC power supply.
3. the Ngatively pressurized sampling apparatus and method selected for use of the present invention have versatility, various substrate materials both be suitable for using, as glass, the capillary array electrophoresis microchip that quartz or high molecular polymer are made can be fit to contain the capillary array electrophoresis microchip of varying number capillary electrophoresis separation unit again.
Description of drawings
Fig. 1. three layers of substrate mask artwork of three-dimensional chip capillary array electrophoresis
Fig. 2. the manufacture craft of three-dimensional chip capillary array electrophoresis and bonding location map
Fig. 3. Ngatively pressurized sampling three-dimensional six swimming lane capillary chip array electrophoresis systems
Among the figure: the three-dimensional six swimming lane glass capillary electrophoretic microchips of 1-, 2-high-voltage DC power supply, 3-Ngatively pressurized sampling device, 3-1 wherein, minipump, 3-2 Dewar bottle, 3-3 electric contact vacuum meter, 3-4 three-way solenoid valve, 3-5 timer, 3-6 interface 4-laser induced fluorescence detector, the wherein charge-coupled pick-up unit of 4-1CCD, the 4-2 color filter, the 4-3 lens, 4-4 laser instrument, 4-5 lens, the 4-6 lens, the 4-7 color filter.
The electrophoretogram that separates six samples in the time of Fig. 4 three-dimensional 6 swimming lane microchip capillary arrays
Embodiment:
Embodiment 1:
The three-dimensional capillary array electrophoresis microchip 1 that provides among the present invention is made by three layers of substrate, and substrate material can be glass, quartz or high molecular polymer.In the boring of the correct position place of upper strata substrate, as the public buffer pool B of three-dimensional capillary array electrophoresis microchip
T, public damping fluid waste liquid pool BW
T, each sample solution pool S
1-S
nOutlet and public sample waste liquid pool SW
TImport.On the substrate of middle one deck, processing contains the channel network of a large amount of capillary electrophoresis separation unit, and each separative element separates swimming lane by one and a sample intake passage is formed.These entrance points that separate swimming lane are by a public damping fluid channel B
BusBe linked together, and pass through at the aperture of substrate corresponding position, upper strata and public buffer pool B
TBe associated in together; The endpiece of separation swimming lane passes through aperture and the damping fluid waste liquid pool BW in the substrate corresponding position on upper strata
TBe connected.The entrance point of every sample intake passage also with each sample solution pool S of substrate corresponding position on upper strata
1-S
nBe connected, simultaneously on bottom substrate, processed a public sample waste fluid channel SW in the corresponding position
BusOn-chip aperture links together with the public sample waste fluid channel that is positioned on the bottom substrate sample intake passage endpiece in each separative element in the middle level by processing, and the relevant position aperture by upper strata and middle level glass substrate, make public sample waste fluid channel that is positioned at bottom substrate and the outlet SW that places the public sample waste liquid of upper strata substrate
TCommunicate.
The mask graph of three-dimensional six swimming lane Capillary Electrophoresis microchips as shown in Figure 1.Wherein, Fig. 1 (a) is the mask pattern on microchip upper strata, and the effect of this piece mask is to determine six sample cell S on the glass substrate of the upper strata of microchip
1-S
6, public buffer pool B
T, public damping fluid waste liquid pool BW
TBore position.The chromium plate that scribbles optical cement need not etching after exposure, directly with removing optical cement after the diamond head boring and the chromium layer can make microchip upper strata substrate.Fig. 1 (b) is the on-chip mask pattern in microchip middle level.The channel network processing that contains a large amount of capillary electrophoresis separation unit is on the substrate of middle level, the effect of this piece mask is the channel network that processing contains 6 capillary electrophoresis separation unit on the glass substrate of chip middle level, and each separative element separates swimming lane by one and a sample intake passage is formed.These preceding 15mm that separate swimming lane become the fan type, to guarantee having enough spaces that sample cell S is installed
1-S
6, the entrance point that separates swimming lane is by a public damping fluid channel B
BusBe linked together, and pass through at the aperture of substrate corresponding position, upper strata and public buffer pool B
TThe back 35mm that separates swimming lane is parallel to each other, spacing is 120 μ L, with the loss that reduces the exciting light energy that laser instrument sends with reduce because the swimming lane overbend makes the degree of separation variation of sample, 6 endpiece that separate swimming lanes pass through aperture and the damping fluid waste liquid pool BW in the substrate corresponding position on upper strata
TBe connected.The entrance point of every sample intake passage also with the aperture and the sample solution pond S of the substrate corresponding position on upper strata
1-S
6Be connected.The chromium plate that scribbles optical cement is through exposure, etching, dechromisation, and obtaining width and be 60 microns, the degree of depth is the channel network of 20 microns three-dimensional six swimming lane glass capillary electrophoretic microchips, public damping fluid channel B
BusWidth be that 1000 microns, the degree of depth are 20 microns.Fig. 1 (c) is the mask pattern of microchip lower floor substrate, and the effect of this piece mask is to determine the public sample waste fluid channel of three-dimensional capillary array electrophoresis microchip SW on the 3rd layer of chip
BusThe position.After photoetching and etching and processing, obtaining width at bottom substrate is that 1000 microns, the degree of depth are 60 microns public sample waste fluid channel.On-chip aperture links together with the public sample waste fluid channel that is positioned on the bottom substrate sample intake passage endpiece in each separative element in the middle level by processing, and the relevant position aperture by upper strata and middle level glass substrate, make public sample waste fluid channel that is positioned at bottom substrate and the outlet (SW that places the public sample waste liquid of upper strata substrate
T) communicate.
After three substrates processing are cleaned, by order shown in Figure 2 with position alignment after method by bonding make their involutions obtain three-dimensional six swimming lane glass Capillary Electrophoresis microchips.Identical in order to guarantee respectively to separate the added electric field intensity of swimming lane two ends when the electrophoretic separation, in public damping fluid channel B
BusIn placed the platinum filament of a diameter 5 microns.Buffer pool B at the upper strata substrate
T, damping fluid waste liquid pool BW
TWith public sample waste liquid pool SW
TPosition, hole place be about 4mm with the bonding internal diameter of epoxy glue respectively, the plastic tube that highly is about 6mm is the corresponding liquid storage tank of conduct respectively, the liquid storage tank volume is about 120 μ L.6 sample cell S
1-6Position, hole place be about 2mm with internal diameter, highly be about each corresponding sample liquid storage tank of conduct of plastic tube of 4mm, the liquid storage tank volume is about 15 μ L.
Embodiment 2: with a plurality of samples of the Ngatively pressurized sampling three-dimensional six parallel separation determinations of swimming lane capillary chip array electrophoresis system
Ngatively pressurized sampling three-dimensional six swimming lane capillary chip array electrophoresis systems as shown in Figure 3, by three-dimensional six swimming lane glass capillary electrophoretic microchips 1, high-voltage DC power supply 2, Ngatively pressurized sampling device 3 and laser induced fluorescence detector 4 are formed, Ngatively pressurized sampling device 3 is referring to patent of invention (number of patent application: 200610048906.6), be made up of minipump 3-1, Dewar bottle 3-2, electric contact vacuum meter 3-3, three-way solenoid valve 3-4, timer 3-5 and interface 3-6.Laser induced fluorescence detector is by the charge-coupled pick-up unit (4-1) that is called for short CCD, and (4-2,4-7), (4-3,4-5 4-6) form color filter for laser instrument (4-4) and lens.
Referring to Fig. 1 and Fig. 2, damping fluid liquid storage tank B on the micro-fluidic chip 1
TWith damping fluid waste liquid liquid storage tank BW
TBetween be 6 and separate swimming lanes, article 6, on-chip aperture links together with the public sample waste fluid channel that is positioned on the bottom substrate sample intake passage endpiece in the middle level by processing, and the relevant position aperture by upper strata and middle level glass substrate, make public sample waste fluid channel that is positioned at bottom substrate and the public sample waste liquid pool SW that places the upper strata substrate
TCommunicate.Referring to Fig. 3, the b port of three-way solenoid valve 3-4 links to each other with interface 3-6 by duct coupling, and interface 3-6 is installed in the public sample waste liquid pool SW of micro-fluidic chip
TAbove, its concrete structure is referring to patent (number of patent application: 200610048906.6 and 200510050457.4).6 sample liquid storage tank S on micro-fluidic chip
1-6In add 5 microlitres three kinds of different sample solutions, wherein S respectively
1And S
2In add 2.0 * 10
-5M rhodamine 123 and 2.0 * 10
-5The mixed solution of M uranin, S
3And S
4In add 4.0 * 10
-5M rhodamine 123 and 4.0 * 10
-5The mixed solution of M uranin, S
5And S
6In add 8.0 * 10
-5M rhodamine 123 and 8.0 * 10
-5The mixed solution of M uranin is at other liquid storage tanks B
T, BW
TAnd SW
TThe borax electrophoretic buffer that adds the 20mmol/L pH 9.2 of 150,150 and 5 microlitres keeps separating swimming lane two ends liquid storage tank B
TAnd BW
TLiquid level identical, sample liquid storage tank S
1-6The height of middle liquid level is less than separating swimming lane two ends liquid storage tank B
TAnd BW
TLiquid level, public sample waste liquid pool SW
TIn liquid level less than liquid storage tank S
1-6The height of middle liquid level.
Operation steps is:
The maximum vacuum of at first setting electric contact vacuum meter is-500mbar that the minimum vacuum degree is-50mbar.Three-way solenoid valve b end and a end are communicated with, and the c end ends.Connect the minipump power supply, make and form negative pressure in the Dewar bottle, vacuum tightness in the Dewar bottle is-50-500mbar, when reaching to set on the vacuum tightness, vacuum tightness in the bottle prescribes a time limit, electric contact vacuum meter is closed the minipump power supply, prescribe a time limit when vacuum tightness in the bottle is lower than to set under the vacuum tightness, electric contact vacuum meter starts minipump, makes bottle interior vacustat in the scope of setting; Laser rays is focused on the check point place that separates on the swimming lane apart from passage right-angled intersection point 30mm, with LASER EXCITED FLUORESCENCE detection electrophoretic separation result.Separating swimming lane B
TEnd applies+the 2000V high voltage BW
TEnd ground connection.
In the sample introduction stage, three-way solenoid valve b end and c end are communicated with, timer picks up counting simultaneously, and Dewar bottle 3-3 is through interface 3-6 and the public sample waste liquid pool SW of micro-fluidic chip
TCommunicate, make public sample waste liquid pool SW
TThe middle negative pressure that forms, sample solution on the micro-fluidic chip in other liquid storage tanks and damping fluid etc. are passed through processing at the on-chip public sample waste fluid channel SW of lower floor under atmospheric effect
BusTo public sample waste liquid pool SW
TFlow, in 0.5 second, form stable sample plug at 6 sample intake passages with the infall that separates swimming lane simultaneously.
Automatically three-way solenoid valve b end-grain cutting is changed to 0.5 second the time with a end by timer and to be communicated with, make three-dimensional chip capillary array electrophoresis system switch to separation phase from the sample introduction stage.Because a of three-way solenoid valve end directly communicates with atmosphere, thereby makes public sample waste liquid pool SW
TCommunicate with atmosphere, pressure differential between it and other liquid pool disappears immediately simultaneously, thereby make and form sample plug at 6 sample intake passages and the infall that separates swimming lane and be added in the electroosmotic flow carry over score that electric field produced that separates on the swimming lane from swimming lane, the beginning electrophoretic separation, write down electrophoretogram simultaneously, article 6, swimming lane separates the electrophoretogram that obtains simultaneously and sees Fig. 4, and the average transit time of rhodamine and uranin is respectively 15.3 and 33.0s, and its relative standard deviation is respectively 3.5 and 1.5%.And the linear relationship of fluorescence intensity and concentration is good in measurement range, and the regression coefficient of the calibration curve of rhodamine and uranin is respectively 0.996 and 0.999.
Claims (6)
1, a kind of Ngatively pressurized sampling three-dimensional chip capillary array electrophoresis system, it is characterized in that: system comprises the three-dimensional capillary array electrophoresis microchip that contains a plurality of capillary electrophoresis separation unit, minipump is as the Ngatively pressurized sampling of negative pressure source and the device that separates, and high-voltage DC power supply and laser fluorescence detection system are formed.Described three-dimensional capillary array electrophoresis microchip is made by three layers of substrate, and public buffer pool B is arranged on the substrate of upper strata
T, public damping fluid waste liquid pool BW
T, each sample solution pool S
1-S
nWith public sample waste liquid pool SW
TThe channel network that a plurality of capillary electrophoresis separation unit is arranged on the substrate of middle one deck, each separative element separates swimming lane by one and a sample intake passage is formed, and the entrance point that separates swimming lane is by a public damping fluid channel B
BusBe linked together, and pass through at the aperture of substrate corresponding position, upper strata and public buffer pool B
TBe associated in together, the endpiece of separation swimming lane passes through aperture and the damping fluid waste liquid pool BW in the substrate corresponding position on upper strata
TBe connected, the entrance point of every sample intake passage also with each sample solution pool S of substrate corresponding position on upper strata
1-S
nBe connected; A public sample waste fluid channel SW is arranged on bottom substrate
BusOn-chip aperture links together with the public sample waste fluid channel that is positioned on the bottom substrate sample intake passage endpiece in each separative element in the middle level by processing, and the relevant position aperture by upper strata and middle level glass substrate, make public sample waste fluid channel that is positioned at bottom substrate and the public sample waste liquid pool SW that places the upper strata substrate
TCommunicate; Described Ngatively pressurized sampling device is by minipump, Dewar bottle, electric contact vacuum meter, three-way solenoid valve, timer, and interface is formed, by interface and public sample waste liquid pool SW
TLink to each other; The positive and negative electrode of described high-voltage DC power supply and public buffer pool B
TWith damping fluid waste liquid pool BW
TIn electrophoresis buffer solution be connected; Described laser fluorescence detection system is made up of charge-coupled pick-up unit, color filter, laser instrument and lens.
2, Ngatively pressurized sampling three-dimensional chip capillary array electrophoresis system according to claim 1 is characterized in that using a micro flow control chip negative pressure sampling device, when three-way solenoid valve b end and the connection of c end, at public sample waste liquid pool SW
TThe middle negative pressure that forms, sample solution on the micro-fluidic chip in other liquid storage tanks and damping fluid are passed through to process in the on-chip public sample waste fluid channel of lower floor to public sample waste liquid pool SW under atmospheric effect
TFlow, in 0.5 second, form stable sample plug at a plurality of sample intake passages with the infall that separates swimming lane simultaneously.
3, Ngatively pressurized sampling three-dimensional chip capillary array electrophoresis system according to claim 1, it is characterized in that described Ngatively pressurized sampling device, automatically three-way solenoid valve is communicated with to switch to the b end and hold with a by b end and c end 0.5 second the time by timer and is communicated with, make public sample waste liquid pool SW
TCommunicate with atmosphere, the pressure differential between it and other liquid pool disappears immediately simultaneously, forms sample plug at a plurality of sample intake passages and the infall that separates swimming lane and is added in the electroosmotic flow carry over score that electric field produced on the separation swimming lane from swimming lane thereby make, and begins electrophoretic separation.
4, the described Ngatively pressurized sampling three-dimensional chip capillary array electrophoresis system of claim 1, it is characterized in that capillary electrophoresis separation swimming lane in the array chip capillary electrophoresis chip and public sample waste fluid channel on two different planes, sample introduction and compartment analysis when using a Ngatively pressurized sampling device just can realize many swimming lanes microchip capillary electrophoresis.
5, Ngatively pressurized sampling three-dimensional chip capillary array electrophoresis system according to claim 1 is characterized in that, in public damping fluid channel B
BusIn placed the platinum filament of 5 microns of diameters, and by at the aperture of substrate corresponding position, upper strata and public buffer pool B
TBe associated in together; The endpiece of separation swimming lane passes through aperture and the damping fluid waste liquid pool BW in the substrate corresponding position on upper strata
TBe connected, with the positive and negative electrode of an adjustable high-voltage DC power supply and public buffer pool B
TWith damping fluid waste liquid pool BW
TIn electrophoresis buffer solution be connected, three-way solenoid valve b end-grain cutting change to a end after, a plurality of sample intake passages and the infall that separates swimming lane form sample plug and are added in the identical electroosmotic flow carry over score that electric field produced that separates on the swimming lane from swimming lane, carry out replicate determination in a plurality of separation swimming lanes simultaneously.
6, according to claim 1 or 4 described Ngatively pressurized sampling three-dimensional chip capillary array electrophoresis systems, it is characterized in that public buffer pool B
T, public damping fluid waste liquid pool BW
T, and public sample waste liquid pool SW
TThe liquid storage tank volume is about 120 μ L, and each sample cell volume is 15 μ L, and the width of the channel network of three-dimensional glass Capillary Electrophoresis microchip is that 60 microns, the degree of depth are 20 microns, public damping fluid channel B
BusWidth be that 1000 microns, the degree of depth are 20 microns, public sample waste fluid channel SW
BusWidth be that 1000 microns, the degree of depth are 60 microns.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB2006100531415A CN100427944C (en) | 2006-08-25 | 2006-08-25 | Ngatively pressurized sampling three-dimensional chip capillary array electrophoresis system |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB2006100531415A CN100427944C (en) | 2006-08-25 | 2006-08-25 | Ngatively pressurized sampling three-dimensional chip capillary array electrophoresis system |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1908647A true CN1908647A (en) | 2007-02-07 |
| CN100427944C CN100427944C (en) | 2008-10-22 |
Family
ID=37699818
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CNB2006100531415A Expired - Fee Related CN100427944C (en) | 2006-08-25 | 2006-08-25 | Ngatively pressurized sampling three-dimensional chip capillary array electrophoresis system |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN100427944C (en) |
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103454221A (en) * | 2013-09-12 | 2013-12-18 | 北京吉天仪器有限公司 | Negative-pressure liquid feeding device based on liquid core wave guide pipe |
| CN103901091A (en) * | 2014-04-22 | 2014-07-02 | 许昌学院 | High-voltage power supply system for capillary electrophoresis chip |
| CN105044052A (en) * | 2015-09-07 | 2015-11-11 | 中国科学院近代物理研究所 | Laser spectrum analysis method and device for elements in liquid |
| CN108593748A (en) * | 2018-01-26 | 2018-09-28 | 南京溯远基因科技有限公司 | capillary and DNA sequencer |
| CN110133772A (en) * | 2019-06-25 | 2019-08-16 | 南京溯远基因科技有限公司 | Aperture device and gene sequencer |
| CN118477710A (en) * | 2024-07-12 | 2024-08-13 | 杭州聚致生物科技有限公司 | Microfluidic electrophoresis chip for DNA separation analysis |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104422780B (en) * | 2013-08-28 | 2016-04-06 | 中国科学院大连化学物理研究所 | A kind of protein express-analysis pick-up unit based on the totally-enclosed system of micro-fluidic chip |
Family Cites Families (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6062261A (en) * | 1998-12-16 | 2000-05-16 | Lockheed Martin Energy Research Corporation | MicrofluIdic circuit designs for performing electrokinetic manipulations that reduce the number of voltage sources and fluid reservoirs |
| AU1438701A (en) * | 1999-10-27 | 2001-05-08 | Caliper Technologies Corporation | Pressure induced reagent introduction and electrophoretic separation |
| US6939452B2 (en) * | 2000-01-18 | 2005-09-06 | Northeastern University | Parallel sample loading and injection device for multichannel microfluidic devices |
| US7037417B2 (en) * | 2001-03-19 | 2006-05-02 | Ecole Polytechnique Federale De Lausanne | Mechanical control of fluids in micro-analytical devices |
| CN2552003Y (en) * | 2002-07-18 | 2003-05-21 | 中国科学院大连化学物理研究所 | Micro fluidic chip |
| CN2727734Y (en) * | 2004-05-19 | 2005-09-21 | 中国科学院大连化学物理研究所 | A array micro-flow controlled chip instrument |
| CN1322004C (en) * | 2004-07-15 | 2007-06-20 | 中国科学院大连化学物理研究所 | Micro-fluidic chip for isolation analysis of protein and isolation analysis method |
| CN100401048C (en) * | 2005-05-19 | 2008-07-09 | 复旦大学 | Multiple channel micro-flow control chip, process for making same and use thereof |
| CN100422731C (en) * | 2005-06-27 | 2008-10-01 | 浙江大学 | Micro flow control chip negative pressure sampling and separating device |
| CN1793890A (en) * | 2006-01-05 | 2006-06-28 | 浙江大学 | Device for microflow controlled chip negatire pressure sample injection and separation using micro vacuum pump |
-
2006
- 2006-08-25 CN CNB2006100531415A patent/CN100427944C/en not_active Expired - Fee Related
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103454221A (en) * | 2013-09-12 | 2013-12-18 | 北京吉天仪器有限公司 | Negative-pressure liquid feeding device based on liquid core wave guide pipe |
| CN103454221B (en) * | 2013-09-12 | 2016-08-31 | 北京吉天仪器有限公司 | Negative pressure feeding device based on liquid core waveguide pipe |
| CN103901091A (en) * | 2014-04-22 | 2014-07-02 | 许昌学院 | High-voltage power supply system for capillary electrophoresis chip |
| CN105044052A (en) * | 2015-09-07 | 2015-11-11 | 中国科学院近代物理研究所 | Laser spectrum analysis method and device for elements in liquid |
| CN108593748A (en) * | 2018-01-26 | 2018-09-28 | 南京溯远基因科技有限公司 | capillary and DNA sequencer |
| CN108593748B (en) * | 2018-01-26 | 2024-04-30 | 南京溯远基因科技有限公司 | Capillary and DNA sequencer |
| CN110133772A (en) * | 2019-06-25 | 2019-08-16 | 南京溯远基因科技有限公司 | Aperture device and gene sequencer |
| CN110133772B (en) * | 2019-06-25 | 2024-06-11 | 南京溯远基因科技有限公司 | Diaphragm device and gene sequencer |
| CN118477710A (en) * | 2024-07-12 | 2024-08-13 | 杭州聚致生物科技有限公司 | Microfluidic electrophoresis chip for DNA separation analysis |
Also Published As
| Publication number | Publication date |
|---|---|
| CN100427944C (en) | 2008-10-22 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| US7419784B2 (en) | Methods, systems and apparatus for separation and isolation of one or more sample components of a sample biological material | |
| CN100427944C (en) | Ngatively pressurized sampling three-dimensional chip capillary array electrophoresis system | |
| KR100852348B1 (en) | Analyte injection system | |
| US6974526B2 (en) | Plastic microfluidics enabling two-dimensional protein separations in proteome analysis | |
| US7153407B2 (en) | Multi-dimensional electrophoresis apparatus | |
| US6991713B2 (en) | Methods and apparatus for processing a sample of biomolecular analyte using a microfabricated device | |
| Long et al. | Integration of nanoporous membranes for sample filtration/preconcentration in microchip electrophoresis | |
| US7641780B2 (en) | Two-dimensional microfluidics for protein separations and gene analysis | |
| US20070090026A1 (en) | Continuous biomolecule separation in a nanofilter | |
| Peng et al. | Recent innovations in protein separation on microchips by electrophoretic methods | |
| JP2007504472A (en) | Analyte injection system | |
| JPH11502618A (en) | Capillary electrophoresis apparatus and method | |
| EP2407777B1 (en) | Extraction of analytes separed by isotachophoresis | |
| US10888863B2 (en) | One-step protein analysis using slanted nanofilter array | |
| WO2002043827A2 (en) | Microstructure apparatus and method for separating differently charged molecules using an applied electric field | |
| EP1044716A1 (en) | Micropreparative isoelectric focussing | |
| WO2004038399A1 (en) | Method of controlling migration of substance | |
| US20030087290A1 (en) | Bio-affinity porous matrix in microfluidic channels | |
| US9409357B1 (en) | Devices, systems, and methods for microscale isoelectric fractionation | |
| Liu et al. | Studying drug–plasma protein interactions by two‐injector microchip electrophoresis frontal analysis | |
| CN2650127Y (en) | Integrated micropump type capillary electrophoresis chip capable of rapid feeding and changing samples | |
| Olson et al. | Experimental study of the effect of disorder on DNA dynamics in post arrays during electrophoresis | |
| US11313830B2 (en) | Electrophoretic separation of biomolecules | |
| CN1540335A (en) | Integrated minityped magnetic pump model capillary electrophoresis chip | |
| CN1779431A (en) | On-line protein electrophoretic preconcentration and electrophoretic separating analysis after preconcentration and special microflow controlled chip |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| C17 | Cessation of patent right | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20081022 Termination date: 20100825 |