CN1869662A - Multi-channel column imaging fluorescent detector - Google Patents
Multi-channel column imaging fluorescent detector Download PDFInfo
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- CN1869662A CN1869662A CNA2006100805218A CN200610080521A CN1869662A CN 1869662 A CN1869662 A CN 1869662A CN A2006100805218 A CNA2006100805218 A CN A2006100805218A CN 200610080521 A CN200610080521 A CN 200610080521A CN 1869662 A CN1869662 A CN 1869662A
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02B—CLIMATE CHANGE MITIGATION TECHNOLOGIES RELATED TO BUILDINGS, e.g. HOUSING, HOUSE APPLIANCES OR RELATED END-USER APPLICATIONS
- Y02B20/00—Energy efficient lighting technologies, e.g. halogen lamps or gas discharge lamps
- Y02B20/30—Semiconductor lamps, e.g. solid state lamps [SSL] light emitting diodes [LED] or organic LED [OLED]
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Abstract
The invention relates to a multichannel column imaging fluorescence detector, comprising: excitation light source, microflow control chip, emitted light filter plate, high voltage power supply and computer control system, in turn arranged, and characterized in that: the excitation light source adopts small-molecular organic LED and plural electrodes of the light source is connected by external circuit so as to make it emit single linear light source or plural different-length linear light sources according to the requirements; the ultrathin excited light filter plate is arranged between the excitation light source and microflow control chip, lens and charge coupling detector are arranged on top of the emitted light filter plate. And the invention can conveniently implement column imaging detection mode without optical fiber, cylindrical prism and complex optical components, further reducing bulk and cost and effectively improving detection sensitivity.
Description
Technical field
The present invention relates to a kind of detecting device,, adopt the column imaging fluorescent detector of Organic Light Emitting Diode (OLED) as the realized multi-channel detection of light source particularly about a kind of micro-fluidic chip isoelectric focusing electrophoresis system that is used for.
Background technology
Protein is a kind of ampholyte molecule, in the pH environment greater than its isoelectric point, can be dissociated into electronegative ion, and electrophoresis is to anodal in electric field; And in pH environment, be dissociated into positively charged ion less than its isoelectric point, to the negative pole swimming, in the pH environment that equals its isoelectric point, be that (isoelectric point pI) just stops net charge, and travel motion and the diffusion motion of charged molecule under electric field action reaches balance at this moment when being zero.If in the environment of a pH gradient, carry out this electrophoresis because the isoelectric point difference of range protein, just can the molecule of different proteins by their isoelectric point concentrated be separated in different district's bands.Isoelectric focusing electrophoresis (isoelectric focusing, IEF) technology is exactly this characteristic according to protein, with polyacrylamide (or other medium) is the electrophoresis holder, and add carrier ampholyte (carrier ampholytes) therein, and under electric field action, protein swimming in the pH gradient gel, when the pH place that migrates to pI, then no longer swimming, and be condensed into narrow district's band, thus realize concentrating and separating of protein example.This method has been widely used in the bidirectional electrophoresis technique in the protein science research.
Capillary isoelectric focusing (capillary isoelectric focusing, cIEF) be on traditional dull and stereotyped IEF basis, to grow up, this technology has overcome the convection problem of dull and stereotyped IEF, can obtain higher separation efficiency, therefore is widely used gradually in recent years.
Different with other electrophoretic, IEF is a kind of balancing technique, and promptly protein electrophorese is just no longer mobile behind the isoelectric point position separately, and system reaches balance, the sample area band can not continue under electric field action through detecting device, therefore has the problem how to detect.The detection method of cIEF technology employing at present mainly contains three kinds: single-point detects, post scanning detects and the post imaging detects.It is exactly to adopt pressure, chemistry or do not eliminate method such as electroosmotic flow that so-called single-point detects, and promotes the sample area band and passes through detecting device and detect.This method can be carried out the cIEF clastotype on general goods Capillary Electrophoresis instrument, needn't carry out the system reform.But also exist to need long analysis time (30～45min), cause problems such as protein sample precipitation and loss separation efficiency easily.It is to finish the back at IEF to move the method for whole kapillary by check point by mechanical hook-up that post scanning detects, and has avoided sample area band moving in kapillary on this engineering philosophy, the problem above having solved.But, can cause that background noise significantly improves when moving, simultaneously owing to remove the broadening that high pressure also can cause the sample area band in the testing process owing to kapillary.It generally is to adopt a branch of optical fiber that light source is drawn that the post imaging detects, and is arranged in line style and converges to kapillary through cylindrical lens, adopts charge-coupled detector(CCD) (CCD) or photodiode array to realize the post imaging then.This technology does not need mobile kapillary or sample area band, and sacrificial separation efficient will not foreshorten in the 5min simultaneously analysis time, but there is the uneven problem of bias light in the general structure more complicated of this system.For this reason, people such as Pawliszyn have been developed a kind of technology based on axial incident imaging, this technology adopts laser (Argon ion laser and semiconductor laser) as light source, scioptics or slit and calibrating installation, make light source from axis of bore to incident, by CCD from receive fluorescence signal perpendicular to the kapillary direction.The optical fiber guiding system has been saved in this design, has reduced the background that causes because of the capillary wall scattered light, has improved the sensitivity of system.But axially incident requires very strict to the optic correction system of kapillary and laser, be difficult for realizing, and because laser cause that by absorption and the scattering of understanding in the kapillary process because of the fluorescent samples band decay to a certain degree takes place the intensity of light source, has limited this system and has been used for detection by quantitative.
The key of above-mentioned post imaging technique is how to convert pointolite to have sufficient intensity uniformly linear light sources.
Summary of the invention
Fundamental purpose of the present invention provides a kind of novel simple and practical column imaging fluorescent detector that is used for cIEF or chip I EF, and volume of the present invention is little, cost is low, and is convenient and practical, and can realize hyperchannel IEF separation simultaneously and the function that detects.
For achieving the above object, the present invention takes following technical scheme: a kind of multi-channel column imaging fluorescent detector, it comprises excitation source, micro-fluidic chip, emission light optical filter, high-voltage power supply and the computer control system that is provided with in regular turn, it is characterized in that: described excitation source adopts the micromolecule Organic Light Emitting Diode, ultra-thin exciting light optical filter is set between described excitation source and described micro-fluidic chip, lens and charge-coupled detector(CCD) is set at the top of described emission light optical filter.
In the technical scheme of the invention described above, described excitation source comprises a glass substrate, be provided with a plurality of indiums-tin-oxide anode on the surface of described glass substrate, be provided with the organism coating that two-layer employing vacuum coating technology plates at described anode surface, constitute the nanoscale hole and import layer and electron transfer layer into, be provided with a plurality of metallic cathodes in described electric transmission laminar surface plating, the cover glass of described cathode surface setting and the encapsulation of described glass substrate is connected with OLED control power supply between each described indium-tin-oxide anode and the metallic cathode.
In the above technical scheme of the present invention, described OLED control power supply is 0～15 volt of adjustable direct supply, to control the linear light sources of each described indium-tin-oxide anode and metallic cathode emission wall scroll or many different lengths.
In the above technical scheme of the present invention, the thickness of described exciting light optical filter is less than 400 μ m.
In the above technical scheme of the present invention, the thickness of described exciting light optical filter is 300 μ m.
In the above technical scheme of the present invention, described light source is 1mm to the distance of described micro-fluidic chip upper channel.
In the above technical scheme of the present invention, the thickness of described micro-fluidic chip egative film is 100 μ m, and described micro-fluidic chip adopts a kind of in glass or the poly-dimethoxy silane of high polymer material, the polymethylmethacrylate.
The present invention is owing to take above technical scheme, and it has the following advantages: 1, the present invention adopts micromolecule OLED as excitation source, replaces original Argon ion laser and semiconductor laser, and the instrument volume is reduced greatly.Needn't adopt optical device and structures such as dichroic prism, lens combination, simplify apparatus structure greatly.2, compare with existing post imaging system, the present invention obtains wall scroll/many illuminating sources by design external circuit control OLED electrod-array, and is simple in structure, is easy to realize.3, to adopt thickness only be ultra-thin exciter filter about 300 μ m in the present invention, on the one hand the parasitic light that covers fluorescence area in the exciting light filtered, and improved the detection sensitivity of instrument effectively; Dwindle the distance between light source and the microchannel on the other hand as far as possible, further dwindled the volume of instrument.4, because the present invention has adopted ultra-thin exciter filter, and needn't adopt the expensive signal processing and amplifying of this class of lock-in amplifier system, cost of the present invention is further reduced.Therefore the present invention greatly reduces the cost and the volume of instrument with the fluorescence detector of Organic Light Emitting Diode (OLED) as light source, simplifies the structure.OLED has the various adjustable characteristic of shape as a kind of light source, promptly can use as a linear light sources, and is convenient and simple, do not need any optical fiber and lens combination, remedied the some shortcomings that exist in the existing post imaging design effectively.
Description of drawings
Fig. 1 is a STRUCTURE DECOMPOSITION synoptic diagram of the present invention
Fig. 2 is an OLED structural representation of the present invention
Fig. 3 is the impact effect figure of exciter filter to the OLED spectral line of emission
Fig. 4 a～Fig. 4 c is the testing result figure of first embodiment of the invention
Fig. 5 a～Fig. 5 c is the testing result figure of second embodiment of the invention
Specific implementation method
The present invention is described in detail below in conjunction with embodiment and conjunction with figs..
As shown in Figure 1, the present invention includes light source 1, exciting light optical filter 2, micro-fluidic chip 3, emission light optical filter 4, camera lens 5, charge-coupled detector(CCD) 6, high-voltage power supply 7, computing machine 8 and OLED control power supply 9.
As shown in Figure 2, the present invention adopts micromolecule OLED as light source 1, it comprises a glass substrate 15 that adopts the conventional criteria photoetching technique to make, glass substrate 15 is provided with a plurality of ITO (indium-tin-oxide) anode 10, each ito anode 10 is provided with the organism coating that two-layer employing vacuum coating technology plates, constitute the nanoscale hole and import layer 11 and electron transfer layer 12 into, upper surface at electron transfer layer 12 also is coated with a plurality of metallic cathodes 13, and OLED control power supply 9 is arranged between each metallic cathode 13 and each ito anode 10.OLED control power supply 9 is 0～15 volt of adjustable direct supply, can buy from the market to obtain, also can be to producer's customization of producing power supply.When ito anode 10 and metallic cathode 13 injected holes with electronics imports layer 11 in the hole and electron transfer layer 12 meets, exciton goes to swash compound tense, has just produced visible light.By regulating the output voltage of OLED control power supply 9, just can make a plurality of ito anodes 10 in oled light source 1 and wall scroll or many linear light sources that metallic cathode 13 is launched different scale as required.Encapsulate with a cover glass 14 at last, constitute the flat light source 1 of OLED.
As shown in Figure 1, the volume of light source 1 size is suitable with micro-fluidic chip 3, and is tabular and is easy to carry out integrated with micro-fluidic chip 3.When 4.5～12V DC voltage of OLED control power supply 9 outputs was added between ito anode 10 and the metallic cathode 13, light source 1 can send the exciting light of respective wavelength and certain intensity, is used for fluoroscopic examination.According to the different organism that plating on the ito anode 10 of light source 1 is established, can make the light source 1 that sends green glow, ruddiness, blue light and ultraviolet light.The present invention replaces original Argon ion laser and semiconductor laser with micromolecule OLED flat-plate light source 1, instrument body is amassed reduce greatly.Because the micromolecule oled light source that the present invention adopts obtains easily, and price is comparatively cheap, promotes the use of so the present invention is beneficial to, and has better practicability.
The present invention is provided with the thick ultra-thin exciting light optical filter 2 of 300 μ m between light source 1 and micro-fluidic chip 3, the parasitic light that covers surveyed area in the exciting light that exciting light optical filter 2 can send light source 1 filters.The egative film thickness of micro-fluidic chip 3 only is 100 μ m, can adopt any transparent matrix material to constitute, as glass or PDMS (poly-dimethoxy silane), PMMA (polymethylmethacrylate) the superpolymer material of etc.ing, and employing standard photoetching and soft lithographic technique are processed into.The top of micro-fluidic chip 3 sets gradually emission light optical filter 4, lens 5 and charge-coupled detector(CCD) 6, after the fluorescence signal that the oled light source 1 passage excited sample band by micro-fluidic chip 3 produces directly filters through emission optical filter 4, receive by lens 5, through charge-coupled detector(CCD) 6 imagings, transfer to computing machine 8 at last, write down the line data of going forward side by side by computing machine 8 and handle.The output port of high-voltage power supply 7 is connected with each sample cell on the micro-fluidic chip 3 respectively, and the com interface of high-voltage power supply 7 other ends is connected with computing machine 8, is used for controlling electrophoresis sample introduction and lock out operation on the micro-fluidic chip 3.
It only is that the thick ultra-thin exciting light optical filter 2 of 300 μ m removes the parasitic light in the excitation source 1 that the present invention is provided with thickness, and as shown in Figure 3, the left side ordinate is the emission light intensity of light source 1 among the figure, and the right side ordinate is the optical filter transmitance.The transmitted spectrum a of exciting light optical filter 2 begins to end from 555nm, and the transmitted spectrum b of emission light optical filter 8 approximately from 560nm begin by.The spectral line of emission that light source 1 is handled without any optical filtering is c, and wherein about 1/4 spectrum covers emitting area, can produce interference by 4 couples of fluoroscopic examination results of emission light optical filter.Add that between light source 1 and micro-fluidic chip 5 exciting light optical filter 2 can filter this part parasitic light, the thickness of exciting light optical filter 2 is got over Bao Yuehao below 400 μ m.Increase the sensitivity that exciting light optical filter 2 can improve system as can be known by the excitation light emission spectral line d after exciting light optical filter 2 filters among the figure.Therefore, exciting light optical filter 2 can filter the parasitic light that covers fluorescence area in the exciting light on the one hand, dwindle the distance of light source and microchannel on the other hand as far as possible, improve the detection sensitivity of system, make that the distance of 3 upper channels is about 1mm from light source 1 to micro-fluidic chip.
Be that example is to being elaborated below with the green-light source.
Example 1: with fluorescence phycoerythrin (R-phycoerythrin) as sample, be disposed in the buffer solution that contains 2% ampholyte (ampholyte) and 1% hydroxypropyl methylcellulose (HPMC), charge into and carry out the isoelectric focusing separation in the micro-fluidic chip, adopt the present invention to detect.Before the operation micro-fluidic chip passage is rinsed well repeatedly with deionized water, be full of the buffer solution that degass through ultrasonic, add the above-mentioned solution of 5 μ L to Fig. 1 sample cell, exerting pressure makes it be full of the microchannel, uses liquid-transfering gun sucking-off surplus solution then.Then add 5 μ L 10mMH in anode and the cathode sample pond respectively
3PO
4With 20mMNaOH solution.Apply 700V/cm electric field intensity between anode and negative electrode, carry out isoelectric focusing electrophoresis.Respectively the electrophoresis spectrogram that obtains of the different time sections in the isoelectric focusing process as shown in Figure 4, wherein the time of a, b, c is respectively to apply the spectrogram that 4.53s, 5.20s and 5.87s write down behind the electric field among Fig. 4.
Example 2: obtain three linear light sources by control oled light source, in micro-fluidic chip, carry out isoelectric focusing electrophoresis with three parallel channels, all the other deposition conditions are with example 1, the result who obtains as shown in Figure 5, wherein a, b, c are respectively the spectrograms that the same concentrations sample obtains after the isoelectric focusing in three passages among Fig. 5.
By above-mentioned example, explanation can be used for micro-fluidic chip isoelectric focusing electrophoresis system with OLED easily as light source, compare with conventional detector and to have characteristics such as volume is little, cost is low, simple in structure, and realize the purpose that hyperchannel detects simultaneously easily.
Claims (11)
1, a kind of multi-channel column imaging fluorescent detector, it comprises excitation source, micro-fluidic chip, emission light optical filter, high-voltage power supply and the computer control system that is provided with in regular turn, it is characterized in that: described excitation source adopts the micromolecule Organic Light Emitting Diode, ultra-thin exciting light optical filter is set between described excitation source and described micro-fluidic chip, lens and charge-coupled detector(CCD) is set at the top of described emission light optical filter.
2, a kind of multi-channel column imaging fluorescent detector as claimed in claim 1, it is characterized in that: described excitation source comprises a glass substrate, be provided with a plurality of indiums-tin-oxide anode on the surface of described glass substrate, be provided with the organism coating that two-layer employing vacuum coating technology plates at described anode surface, constitute the nanoscale hole and import layer and electron transfer layer into, be provided with a plurality of metallic cathodes in described electric transmission laminar surface plating, the cover glass of described cathode surface setting and the encapsulation of described glass substrate is connected with OLED control power supply between each described indium-tin-oxide anode and the metallic cathode.
3, a kind of multi-channel column imaging fluorescent detector as claimed in claim 1, it is characterized in that: described OLED control power supply is 0～15 volt of adjustable direct supply, to control the linear light sources of each described indium-tin-oxide anode and metallic cathode emission wall scroll or many different lengths.
3, a kind of multi-channel column imaging fluorescent detector as claimed in claim 1 is characterized in that: the thickness of described exciting light optical filter is less than 400 μ m.
4, a kind of multi-channel column imaging fluorescent detector as claimed in claim 2 is characterized in that: the thickness of described exciting light optical filter is less than 400 μ m.
5, as claim 3 or 4 described a kind of multi-channel column imaging fluorescent detectors, it is characterized in that: the thickness of described exciting light optical filter is 300 μ m.
6, as claim 1 or 2 or 3 or 4 described a kind of multi-channel column imaging fluorescent detectors, it is characterized in that: described light source is 1mm to the distance of described micro-fluidic chip upper channel.
7, a kind of multi-channel column imaging fluorescent detector as claimed in claim 5 is characterized in that: described light source is 1mm to the distance of described micro-fluidic chip passage.
8, as claim 1 or 2 or 3 or 4 or 7 described a kind of multi-channel column imaging fluorescent detectors, it is characterized in that: the thickness of described micro-fluidic chip egative film is 100 μ m, and described micro-fluidic chip adopts a kind of in glass or the poly-dimethoxy silane of high polymer material, the polymethylmethacrylate.
9, a kind of multi-channel column imaging fluorescent detector as claimed in claim 5, it is characterized in that: the thickness of described micro-fluidic chip egative film is 100 μ m, and described micro-fluidic chip adopts a kind of in glass or the poly-dimethoxy silane of high polymer material, the polymethylmethacrylate.
10, a kind of multi-channel column imaging fluorescent detector as claimed in claim 6, it is characterized in that: the thickness of described micro-fluidic chip egative film is 100 μ m, and described micro-fluidic chip adopts a kind of in glass or the poly-dimethoxy silane of high polymer material, the polymethylmethacrylate.
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| CN102305778A (en) * | 2011-05-17 | 2012-01-04 | 易定容 | Micro-multispectral fluorescence reception and treatment system |
| CN102507523A (en) * | 2011-11-09 | 2012-06-20 | 大连理工大学 | OLED (Organic Light-Emitting Diode)- and OPD (Organic Photodetector)-based cascading vertical integration capillary electrophoresis chip |
| CN102628805A (en) * | 2012-04-26 | 2012-08-08 | 大连理工大学 | Microfluidic chip fluorescence detection system based on photonic crystal filter plates |
| CN103308587A (en) * | 2013-05-30 | 2013-09-18 | 上海大学 | Hollow fiber electrophoresis gel imaging system |
| CN103409317A (en) * | 2013-07-23 | 2013-11-27 | 广州市第一人民医院 | Capillary bioanalysis system, and analytical method and applications thereof |
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2006
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| CN102305778A (en) * | 2011-05-17 | 2012-01-04 | 易定容 | Micro-multispectral fluorescence reception and treatment system |
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