CN1865932A - Fluorometric detector for micro-fluidic chip system - Google Patents
Fluorometric detector for micro-fluidic chip system Download PDFInfo
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- CN1865932A CN1865932A CN 200510070976 CN200510070976A CN1865932A CN 1865932 A CN1865932 A CN 1865932A CN 200510070976 CN200510070976 CN 200510070976 CN 200510070976 A CN200510070976 A CN 200510070976A CN 1865932 A CN1865932 A CN 1865932A
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Abstract
The related fluorescence detector for micro flow control chip system comprises: in turns, a laser source used small molecular OLED, a pin hole, a substrate, a micro flow control chip, a ultra-thin filter with a photomultiplier on top for emission light between the source and hole, a high-voltage power, and a computer control system. This invention improves detection sensitivity, and reduces cost.
Description
Technical field
The present invention relates to a kind of pick-up unit, particularly about the fluorescence detection device that be used for micro-fluidic chip system of a kind of employing Organic Light Emitting Diode (OLED) as light source.
Background technology
Along with progress of science and technology, analytical instrument and analysis science are also just experiencing deep change, one of them day by day obvious development trend be exactly the microminiaturization of chemical analysis apparatus, integrated and portability.Micro-total analysis system (Micro total analysis systems, the notion of μ-TAS) is to propose the beginning of the nineties, its objective is microminiaturization by chemical analysis apparatus and integrated, to greatest extent the function of assay laboratory is transferred in the portable analytical equipment, even be integrated on the chip of heart size, therefore be also referred to as chip lab (Lab-on-a-chip), one of technical barrier wherein is exactly how to comprise that many equipment of detecting device is microminiaturized, integrated in the lump.
At present, the detecting device that is used for micro-fluidic chip mainly contains laser-induced fluorescence (LIF) (LIF), galvanochemistry, chemiluminescence, molecular spectrum, mass spectrum etc.Wherein laser-Induced Fluorescence Detection is to use in microfluidic system the earliest, and still uses more detection means so far, also is unique adopted detecting device in commercialization micro-fluidic chip analytic system at present.It is bulky that but traditional LIF detection system exists usually, costs an arm and a leg, and the shortcoming that power consumption is higher is not suitable for micro-fluidic chip system.
For develop volume little, be easy to carry, the cheap LIF detecting device that is used for micro-fluidic chip system, domestic Mr. Fang Zhaolun of Northeastern University leader's research group adopts the 635nm small semiconductor laser as excitation source, volume and cost have been reduced to a great extent, but owing to equally with common LIF system need to add optical elements such as prism, lens between light source and detecting device, distance is integrated to still have suitable distance.Semiconductor light-emitting-diode (LED) is another kind of excitation source, and its volume is littler, and price is cheaper, can simplify the structure of detecting device, reduces the cost of detecting device.Researchists such as Burns have made the silicon photoelectric diode photodetector on monocrystalline silicon piece, and plate filter coating, adopt blue LED as excitation source dna material to be carried out fluoroscopic examination.Webster etc. have also reported a kind of LED detection technique of fluorescence that is integrated on the chip, have realized the separation detection of DNA restricted fragment.Though above-mentioned micro-fluidic chip has reduced the volume of system to a great extent, still be not integrated in the chip system as the light emitting diode of light source, but also have the machining precision height, complex process, the shortcoming that cost is higher.
Organic Light Emitting Diode (OLED) is based on a kind of current mode light emitting semiconductor device of organic material, be mainly used in fields such as screen display, pilot lamp, instrument and meter at present, different with the inorganic thin-film electrofluorescence device, OLED mainly depends on the excitonic luminescence that forms in the luminescent layer.The electron hole pair that exciton carries the baby, meeting from ito anode and metal electrode injected holes and electronics respectively forms.When exciton goes to swash compound tense, will produce visible light.According to the difference of luminescent material, OLED can be divided into three classes again: micromolecule OLED, polymkeric substance OLED (being also referred to as PLED) and group of the lanthanides organic metal OLED (also being rare earth OLED).The research work that OLED is used for the detecting device of analytical instrument only just begins, and the relevant report DeMello etc. that has only in April, 2004 London Royal Academy is the work of the miniflow fluorescence detecting system of light source in Lab on a chip above-reported with polymkeric substance OLED therewith.The about 40 μ m of poly-fluorenyl PLED structure light source area * 1000 μ m that they make, maximum emission wavelength is 488nm, and the PLED light source is close to micro-fluidic chip, does not adopt any optical element, and light source distance microchannel distance is 2mm.The fluorescence signal that excites generation is sent in the avalanche diode detecting device through emission light optical filter by a simple confocal system of cover (comprising a pin hole and microcobjective) and is converted to electric signal, again by lock-in amplifier with the signal processing and amplifying, get off by computer recording.They are with this system's separation and detected fluorescein and two kinds of fluorescent dyes of 5-Fluoresceincarboxylic acid.This system is not because to the exciting light processing that filters, the spectrum that light source sends compares broad, the spuious exciting light of a part can pass through detecting device, influenced detection sensitivity to a great extent, have to adopt the expensive signal processing and amplifying of this class of lock-in amplifier system for the sensitivity that improves system, make the with high costs of this system.In addition, the employed poly-fluorenyl PLED light source of this system not only costs an arm and a leg, and the external world can't obtain, and makes this system to promote the use of.
Summary of the invention
Fundamental purpose of the present invention provides a kind of apparatus structure of simplifying, and reduces the instrument volume, reduces cost, and can further improve the fluorescence detection device that is used for micro-fluidic chip system of system's detection sensitivity.
For achieving the above object, the present invention takes following technical scheme: a kind of fluorescence detection device that is used for micro-fluidic chip system, it comprises the excitation source that is provided with in regular turn, pin hole, the micro-fluidic chip egative film, micro-fluidic chip, emission light optical filter, high-voltage power supply and computer control system, 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 pin hole, between described micro-fluidic chip and emission light optical filter, the collar and the optical fiber that is arranged in the described collar are set, photomultiplier are set at the top of described emission light optical filter.
Described excitation source comprises a glass substrate, be provided with indium-tin-oxide anode on the surface of described glass substrate, establish the organism that formation nanoscale hole is imported layer and electron transfer layer in described anode surface plating, establish metallic cathode in described electric transmission laminar surface plating, the cover glass of described cathode surface setting and the encapsulation of described glass substrate.
The thickness of described exciting light optical filter is less than 400 μ m.
The thickness of described exciting light optical filter is 300 μ m.
Described light source is 1mm to the distance of described micro-fluidic chip upper channel.
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 P LED system, it only is ultra-thin exciter filter about 300 μ m that the present invention has adopted thickness, on the one hand the parasitic light that covers fluorescence area in the exciting light is filtered, and has 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.3, 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.
Description of drawings
Fig. 1 is a STRUCTURE DECOMPOSITION synoptic diagram of the present invention
Fig. 2 is the diagrammatic cross-section after the present invention installs
Fig. 3 is the structural representation of the Organic Light Emitting Diode (OLED) that uses of the present invention
Fig. 4 is the impact effect figure of exciter filter to the OLED spectral line of emission
Fig. 5 is the testing result figure of first embodiment of the invention
Fig. 6 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 and Figure 2, the present invention includes light source 1, exciting light optical filter 2, pin hole 3, micro-fluidic chip egative film 4, micro-fluidic chip 5, the collar 6, optical fiber 7, optical filter 8, photomultiplier 9, high-voltage power supply 10 and computing machine 11.
As shown in Figure 3, the present invention adopts micromolecule OLED as light source 1, it comprises a glass substrate 12 that adopts standard photolithography techniques to make, glass substrate 12 is provided with ito anode 13, on ito anode 13, be coated with the organism coating that two-layer employing vacuum coating technology plates, constitute nano level hole and import layer 14 and electron transfer layer 15 into, also plate at the upper surface of electron transfer layer 15 and to be provided with metallic cathode 16, between metallic cathode 16 and ito anode 13, be provided with an adjustable direct supply DC power supply, when ito anode 13 and metallic cathode 16 injected holes with electronics imports layer 14 in the hole and electron transfer layer 15 meets, exciton goes to swash compound tense, has just produced visible light.Encapsulate with a cover glass 17 at last, constitute the flat light source 1 of OLED.The volume of light source 1 size is suitable with micro-fluidic chip 5, and is tabular and is easy to carry out integrated with micro-fluidic chip 5.When 4.5～12V DC voltage of adjustable direct supply output was added between ito anode 13 and the metallic cathode 16, 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 13 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.
As shown in Figure 1, the present invention is provided with the pin hole 3 of thick ultra-thin exciting light optical filter 2 of 300 μ m and thickness of slab 12 μ m in regular turn between light source 1 and micro-fluidic chip egative film 4, the parasitic light that covers surveyed area in the exciting light that exciting light optical filter 2 can send light source 1 filters, and 3 of pin holes can limit the spot size of light source 1 on the passage of micro-fluidic chip 5.The thickness of micro-fluidic chip egative film 4 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 5 is provided with optical fiber 7, and optical fiber 7 is aimed at the passage on the micro-fluidic chip 5 by the collar 6 and is fixed in the above.The other end at optical fiber 7 is provided with emission light optical filter 8 and photomultiplier 9, the fluorescence signal of the excitation light generation of process micro-fluidic chip 5 is received by photomultiplier 9 after emission optical filter 8 filters by optical fiber 7 and amplifies, transfer to computing machine 11 at last, write down the line data of going forward side by side by computing machine 11 and handle.Four road output ports of high-voltage power supply 10 respectively with micro-fluidic chip 5 on sample cell with the buffering solution pool be connected, the com interface of high-voltage power supply 10 other ends is connected with computing machine 11, is used for controlling electrophoresis sample introduction and lock out operation on the micro-fluidic chip 5.
It only is that the thick ultra-thin exciting light optical filter 2 of 300 μ m removes the parasitic light (as shown in Figure 4) in the excitation source 1 that the present invention is provided with thickness, and 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 8 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 5 upper channels has only 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 the Alexa532 dyestuff of variable concentrations as sample, with 4% linear polyacrylamide (LPA), 1 * TBE (89mM Tris, 89mM boric acid, 2mM EDTA pH8.3) is buffer solution, electrophoretic separation in micro-fluidic chip, adopt the present invention to detect, to examine or check sensitivity of the present invention.Before the operation micro-fluidic chip passage is rinsed well repeatedly with deionized water, be full of the buffer solution that degass through ultrasonic, buffer solution pond B in Fig. 1, add 5 μ L buffer solution respectively among buffering waste liquid pool BW and the sample waste liquid pool SW, the sample solution that in sample cell S, adds same amount, sample introduction, separation and detection under reverse voltage, its electric field intensity is respectively 250V/cm and 470V/cm, the testing result that obtains (as shown in Figure 5).For the Alexa532 dyestuff, the minimal detectable concentration that obtains is 7 μ M, and the about 0.7nL fluorescence signal intensity of sampling volume obtains the better linearity relation from 7 μ M to 280 μ M with the concentration increase, departs from the phenomenon of lambert-Bill's curve when concentration continues to increase.
Example 2: the present invention is used on the micro-fluidic chip electrophoretic separation and detects protein example.To 45 μ L concentration is to add 5 μ L10mg/mLAlexa532 dyestuffs in the 20mg/mL bovine serum albumin(BSA) (BSA), and concussion is 1 hour under the room temperature, and 4 ℃ of following reactions are spent the night.Albumen with the above-mentioned Alexa532 mark not doing to be further purified is sample, after 10 times of buffer solution dilutions, carries out electrophoretic separation and adopt this system to detect on micro-fluidic chip, the electrophoretic separation spectrogram (as shown in Figure 6) that obtains.Wherein two electrophoresis peaks, front e, f are unreacted Alexa532 dyestuff spectrum peaks, and latter two electrophoresis peak j, h are that BSA combines product with dyestuff, and deposition condition is consistent with example 1 with buffering solution.
By above-mentioned example, explanation is the separation detection that the micro-fluidic chip fluorescence detecting system of the microminiaturization of light source not only can be used to biological sample as laser induced fluorescence detector with OLED, and the present invention does not adopt optical device and structures such as dichroic prism, lens combination, simplified apparatus structure greatly, volume of the present invention is reduced, realized the microminiaturization of detection system, and can improve the detection sensitivity of system and reduce cost.
Claims (10)
1, a kind of fluorescence detection device that is used for micro-fluidic chip system, it comprises the excitation source that is provided with in regular turn, pin hole, the micro-fluidic chip egative film, micro-fluidic chip, emission light optical filter, high-voltage power supply and computer control system, 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 pin hole, between described micro-fluidic chip and emission light optical filter, the collar and the optical fiber that is arranged in the described collar are set, photomultiplier are set at the top of described emission light optical filter.
2, the fluorescence detection device that is used for micro-fluidic chip system as claimed in claim 1, it is characterized in that: described excitation source comprises a glass substrate, be provided with indium-tin-oxide anode on the surface of described glass substrate, establish the organism that formation nanoscale hole is imported layer and electron transfer layer in described anode surface plating, establish metallic cathode in described electric transmission laminar surface plating, the cover glass of described cathode surface setting and the encapsulation of described glass substrate.
3, the fluorescence detection device that is used for micro-fluidic chip system as claimed in claim 1 is characterized in that: the thickness of described exciting light optical filter is less than 400 μ m.
4, the fluorescence detection device that is used for micro-fluidic chip system 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 being used for the fluorescence detection device of micro-fluidic chip system as described in claim 3 or 4, it is characterized in that: the thickness of described exciting light optical filter is 300 μ m.
6, as claim 1 or 2 or the 3 or 4 described fluorescence detection devices that are used for micro-fluidic chip system, it is characterized in that: described light source is 1mm to the distance of described micro-fluidic chip upper channel.
7, as being used for the fluorescence detection device of micro-fluidic chip system as described in the claim 5, it 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 the 4 or 7 described fluorescence detection devices that are used for micro-fluidic chip system, 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, the fluorescence detection device that is used for micro-fluidic chip system 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, the fluorescence detection device that is used for micro-fluidic chip system 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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| CN105492888A (en) * | 2013-09-05 | 2016-04-13 | 英派尔科技开发有限公司 | Cell culturing and tracking with OLED arrays |
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| CN103134780A (en) * | 2011-12-01 | 2013-06-05 | 中国科学院大连化学物理研究所 | Closely attached type light emitting diode induction fluorescence detector for exciting light path |
| CN103308587A (en) * | 2013-05-30 | 2013-09-18 | 上海大学 | Hollow fiber electrophoresis gel imaging system |
| CN103323439B (en) * | 2013-06-09 | 2016-03-30 | 华侨大学 | A kind of micro-fluidic chip fluorescence excitation device, micro-fluidic chip and preparation method thereof |
| CN103323439A (en) * | 2013-06-09 | 2013-09-25 | 华侨大学 | Micro-fluidic chip fluorescence excitation device, micro-fluidic chip and preparation method thereof |
| CN105492888A (en) * | 2013-09-05 | 2016-04-13 | 英派尔科技开发有限公司 | Cell culturing and tracking with OLED arrays |
| CN103604785A (en) * | 2013-11-05 | 2014-02-26 | 浙江工业大学 | Fluorescence detection system |
| US10114008B2 (en) | 2014-01-21 | 2018-10-30 | Empire Technology Development Llc | Methods and devices for high throughput screening of conditions affecting stem cell differentiation |
| CN106092994A (en) * | 2016-07-27 | 2016-11-09 | 福州大学 | A kind of micro-array chip fluorescence detection method of great power LED |
| CN106092994B (en) * | 2016-07-27 | 2019-05-10 | 福州大学 | A kind of micro-array chip fluorescence detection method of great power LED |
| CN112903793A (en) * | 2021-01-21 | 2021-06-04 | 中国科学技术大学 | Multi-parameter rapid detection device and method for water body |
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