CN101153869A - Device for high-throughout monitoring micro-array biomolecule reaction by light reflection difference method - Google Patents
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- 238000012544 monitoring process Methods 0.000 title claims abstract description 33
- 238000002493 microarray Methods 0.000 title claims abstract description 23
- 238000006243 chemical reaction Methods 0.000 title claims abstract description 20
- 238000000034 method Methods 0.000 title claims abstract description 19
- 229910052710 silicon Inorganic materials 0.000 claims abstract description 24
- 239000010703 silicon Substances 0.000 claims abstract description 24
- XUIMIQQOPSSXEZ-UHFFFAOYSA-N Silicon Chemical compound [Si] XUIMIQQOPSSXEZ-UHFFFAOYSA-N 0.000 claims abstract description 23
- 238000012545 processing Methods 0.000 claims abstract description 22
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- 238000001215 fluorescent labelling Methods 0.000 description 3
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Abstract
The invention provides a device for high-flux monitoring of micro array biomolecular reaction through light reflex difference method, comprising an incident light path, a sample table, an emergent light path and a data acquisition and processing system, wherein, the incident light path comprises a laser, a polarizer, a photoelastic modulator, a phase shifter and a beam expander arranged between the phase shifter and the sample table; the emergent light path comprises a polarization analyzer and an electrooptical signal converter; the electrooptical signal converter comprises a silicon photodiode array, a pulse switching circuit, a first phase-locked amplifier and a second phase-locked amplifier; the output end of the silicon photodiode array is respectively in electric connection with the input ends of the pulse switching circuit, the first phase-locked amplifier and the second phase-locked amplifier. With a simple structure and easy operation, the device reduces the noise caused by moving a sample and greatly improves signal-noise ratio; meanwhile, the device can complete non-mark quick high-flux monitoring of the reactions of the array elements of a micro array at the same time, thereby greatly increasing monitoring efficiency.
Description
Technical Field
The invention relates to a monitoring device for micro-array biomolecule reaction, in particular to a device for high-throughput rapid unmarked monitoring of biomolecule reaction.
Background
Common methods for monitoring biomolecules are fluorescence labeling and surface plasmon resonance. The fluorescence labeling is a common destructive monitoring method, and the method has the defects of complex monitoring process, capability of changing the structure of protein molecules, generation of photobleaching phenomenon and the like; surface plasmon resonance is a commonly used method for monitoring biomolecular interaction without labels at present, and the method requires that a biomolecule microarray is formed on a functional gold film, and has high requirements on the thickness and uniformity of the gold film.
Oblique incidence light reflection difference (OIRD) is a high-sensitivity special optical measurement technique for measuring the optical properties of a surface film, and can monitor the relative reflectivity delta R/R (1 × 10) in the film growth process-5And the change of the coverage angle delta theta is 0.02, and the method is widely applied to the real-time in-situ noninvasive monitoring of the growth process of the thin film material. In recent years, this method has also been applied to monitoring of biomolecules, and as described in document 1(Detection of biomolecular micro arrays with a light emitting fluorescent-labeling agents, SPIE 5328(121), 2004), a light spot having a diameter of 3 μm is used to irradiate a sample in an experiment, a silicon photodiode is used to receive the reflected light, the light path is fixed during the experiment, and the sample to be measured is moved in two dimensions. The whole device comprises an incident light path system consisting of a P-polarized laser light source, a photoelastic modulator, a Pockels cell and a lens which are sequentially arranged on a light path, an emergent light path part consisting of the lens, an analyzer and a photoelectric detector and a data acquisition and processing system, wherein the device is adopted in the literature to carry out biomolecule array synthesis reaction detection, quantitative detection of optical properties of a microarray and detection of the microarray in an aqueous solution under the condition of no fluorescent label. The monitoring device needs long time for monitoring the multi-array element biomolecule microarray reaction, and the movement of the sample brings inevitable system noise to the monitoring result.
The invention content is as follows:
the invention aims to overcome the defects of the existing OIRD biomolecule monitoring technology, and provides a device for monitoring the reaction of microarray biomolecules at high flux by a light reflection difference method, wherein the device does not need to move a sample, has short monitoring time and low system noise.
The invention provides a device for monitoring micro-array biomolecule reaction with high flux by a light reflection difference method, which comprises an incident light path, a sample stage, an emergent light path and a data acquisition and processing system, wherein the incident light path comprises a laser, a polarizer, a photoelastic modulator and a phase shifter, wherein the polarizer, the photoelastic modulator and the phase shifter are sequentially arranged on the light path in front of the output light of the laser; the emergent light path comprises an analyzer and a photoelectric signal converter, the analyzer and the photoelectric signal converter are sequentially arranged in front of an emergent light beam reflected by a sample on the sample stage, the photoelectric signal converter is electrically connected to the data acquisition and processing system, and the device is characterized in that,
the incident light path also comprises a beam expanding device arranged between the phase shifter and the sample stage;
the mechanical joint is composed of two vertical plates vertically fixed on the sample table, an aluminum alloy flat plate for receiving incident light and an aluminum alloy flat plate for reflecting emergent light, wherein the two aluminum alloy flat plates and the two vertical plates are provided with arc-shaped fixing screw holes, the two aluminum alloy flat plates are obliquely fixed on the vertical plates through a bolt, the screw holes are in a quarter arc shape, and the aluminum alloy plate can be adjusted between 0 and 90 degrees;
the incident light path is arranged on the aluminum alloy flat plate for receiving incident light, and the emergent light path is arranged on the aluminum alloy flat plate for reflecting emergent light;
the photoelectric signal converter comprises a silicon photodiode array, a switching circuit, a first phase-locked amplifier and a second phase-locked amplifier, wherein the output end of the silicon photodiode array is electrically connected to the input ends of the switching circuit, the first phase-locked amplifier and the second phase-locked amplifier respectively.
In the above technical solution, the data acquisition and processing system includes a BNC adapter, a data acquisition card and a data processing device; wherein
The BNC adapter receives the output signals of the first phase-locked amplifier and the second phase-locked amplifier, performs channel division processing on the output signals of the phase-locked amplifiers, and divides the output signals of each phase-locked amplifier into two parts, namely amplitude information and phase information;
the data acquisition card acquires data output by the BNC adapter and transmits the data to the data processing device;
and the data processing device stores, analyzes and processes the data sent by the data acquisition card.
In the above technical solution, the beam expanding device is a beam expanding lens group, the beam expanding lens group includes a collimating lens barrel and two continuous zoom beam expanding lenses respectively disposed at two ends of the collimating lens barrel, and the diameter of the cross section of the outgoing beam can be adjusted by adjusting the length of the collimating lens barrel.
In the above technical solution, the incident light path and the emergent light path are connected by a mechanical joint capable of adjusting the incident angle of the light beam.
In the above technical solution, the laser is a linearly polarized He — Ne laser having a wavelength of 632.8 nm.
In the above technical solution, the photoelastic modulator has a modulation frequency Ω of 50 kHz.
In the above technical scheme, the sample stage is a stainless steel platform with a sample fixing mechanism and three-dimensional manual adjustment.
The invention has the advantages that:
(1) the invention adopts the silicon photodiode array as a receiving element of reflected light, when an incident laser beam passes through a series of optical systems and then is incident on a microarray sample as a wide beam, the reflected light is received by the silicon photodiode array after passing through an analyzer, when the incident angle is fixed, the silicon photodiode array can receive all the light reflected by the surface of the biological sample only by adopting the silicon photodiode array without two-dimensional movement of a sample stage, thus not only reducing the noise caused by moving the sample, but also greatly improving the signal-to-noise ratio;
(2) the wide light beam incidence and the silicon photodiode array are adopted to receive the reflected light, so that the device can simultaneously carry out unmarked, rapid and high-flux monitoring on various reactions of a plurality of array elements of the microarray, and the monitoring efficiency is greatly improved;
(3) the device has simple structure, easy operation, high flux and high working efficiency, and can quickly realize the monitoring of the biomolecular reaction without any marking reagent;
(4) the devices used by the invention can be purchased in the market and are easy to realize;
(5) in addition, the incident light path system and the emergent light path system are connected by adopting a mechanical joint, so that the incident angle of the light beam can be randomly adjusted between 0 and 90 degrees.
Drawings
FIG. 1 is a schematic diagram of the apparatus for high-throughput monitoring of the reaction of microarray biomolecules by light reflectance difference method according to the present invention.
Reference numerals:
1-a laser; 2-a polarizer; 3-a photoelastic modulator; 4-a phase shifter;
5-a beam expanding device; 6-sample stage; 7-analyzer; 8-a switching circuit;
a 9-silicon photodiode; 10-a first phase-locked amplifier; 11-a second lock-in amplifier;
12-data acquisition and processing system.
Detailed Description
The invention will be further explained and illustrated with reference to specific examples.
Example 1
Referring to FIG. 1, a device for high throughput monitoring of the reaction of microarray biomolecules by light reflectance contrast method is fabricated.
The device comprises an incident light path, a sample stage 6, an emergent light path and a data acquisition and processing system, wherein the incident light path comprises a polarizer 2, a photoelastic modulator 3, a phase shifter 4 and a beam expanding device 5 which are sequentially arranged on the light path in front of the output light of a laser 1; the emergent light path is divided by an analyzer 7 and a photoelectric signal converter 9, an incident light beam emitted from an incident light path is reflected by a sample on a sample stage 6 and then sequentially passes through the analyzer 7 and the photoelectric signal converter, and the photoelectric signal converter is electrically connected to the data acquisition and processing system 12. The photoelectric signal converter consists of a silicon photodiode array 9, a switching circuit, a first phase-locked amplifier 10 and a second phase-locked amplifier 11, wherein the output end of the silicon photodiode array 9 is electrically connected to the input ends of the switching circuit, the first phase-locked amplifier 10 and the second phase-locked amplifier 11 respectively.
Still adopt mechanical joint to adjust incidence and emergent light in the device of this embodiment, this mechanical joint by two vertical fixation on the sample bench riser, and one accept the aluminum alloy flat board of incident light, with one be used for reflecting the aluminum alloy flat board of emergent light, two aluminum alloy flat boards and two risers on open and to have convex fixed screw, constitute on the riser through a bolt with two dull and stereotyped slants of aluminum alloy, the screw be that the fourth is circular-arc, the aluminum alloy plate can mediate between 0-90, such mediation realization is to the regulation of incidence and emergent light. The incident light path is arranged on an aluminum alloy flat plate for receiving incident light, and the emergent light path is arranged on an aluminum alloy flat plate for reflecting emergent light.
The device of this embodiment uses a He — Ne laser having a power of 4mW and a wavelength of 632.8nm as a light source, and has an exit aperture of 3mm, and a laser beam emitted from the laser 1 is changed into p-polarized light having a polarization direction parallel to an incident surface after the polarization direction is corrected by a polarizer (New Focus5524) 2; then passes through a photoelastic modulator (Hinds Instrument PEM 90)TM)3, generating periodic modulation between p and s polarization states, wherein the modulation frequency is 50 KHz; the polarized modulated light emitted from the photoelastic modulator 3 is introduced into adjustable phase compensation between p and s polarization states through a phase shifter 4 consisting of a quartz multistage half-wave plate with the diameter of 25.4 mm; then a beam broadening lens group serving as a beam expanding device 5 widens a laser beam and then emits the laser beam onto a sample stage 6, wherein the beam broadening lens group consists of a collimating lens barrel and two continuous variable-magnification beam expanding lenses respectively arranged at two ends of the collimating lens barrel, the length of the collimating lens barrel is adjustable, the total length of the collimating lens barrel is maximally 134mm, the applicable wavelength range of the beam broadening lens group is 450-680nm, the beam broadening ratio is 5X-6X, and the diameters of an incident light spot and an emergent light spot are respectively 4mm and 24 mm; the sample table 6 adopts a stainless steel table top purchased in the market and a platform capable of realizing three-dimensional manual adjustment, the adjustment precision is 0.01mm, and the three-dimensional stroke is 50 mm;
the reflected light reflected by the surface of the sample is received by a silicon photodiode array 9 after being regulated by an analyzing analyzer 7(CVI Laser CPAD-10.0-425-675); the first phase-locked amplifier 10 and the second phase-locked amplifier 11(Stanford Research Systems SR830DSP) are respectively electrically connected to the silicon photodiode array 9, and the silicon photodiode array 9 has a photosensitive area of 1.0mm2The square high-speed sensing silicon photodiodes (BPX-65) form a 30 x 30 two-dimensional array. Since the silicon photodiode array 9 has a plurality of silicon photodiodes, in order to make the optical signal received by each silicon photodiode be able to be analyzed by the first phase-locked amplifier 10 and the second phase-locked amplifier 11, a switch circuit is electrically connected to the silicon photodiode array 9, the switch circuit in this embodiment is a relay switching circuit 8, so that the optical signal received by each silicon photodiode is converted into an optical signalThe primary harmonic frequency and the secondary harmonic frequency after being converted into the electric signal can be read by a data acquisition and processing system in sequence;
the data acquisition and processing system 12 in the device adopts a computer, the computer is provided with a data acquisition card (PCI-6220), a BNC adapter (BNC-2110) and an acquisition program written by LabVIEW, electric signals output by a first phase-locked amplifier 10 and a second phase-locked amplifier 11 are firstly processed by channels through the BNC adapter, the signal output by each phase-locked amplifier is divided into two parts of amplitude information and phase information, then the amplitude information and the phase information of each phase-locked amplifier are respectively sent to the data acquisition card for acquisition, and finally the acquired data are transmitted to the data processing program written by the LabView for storage, analysis and processing.
When the device of the invention is adopted, the monitoring precision is 2um, the monitoring of the biological samples of 20 multiplied by 20 to 200 multiplied by 200 biochip arrays can be completed within 2 minutes, wherein the diameter of the array elements is 100um, the distance is 100um, and the monitoring sensitivity reaches 2 multiplied by 10-6rad。
Example 2
In this embodiment, the phase shifter 4 is a pock cell of Cleveland Crystal IMPACT10 type, and the device sensitivity of this embodiment can be improved by 1 order of magnitude as compared with embodiment 1, which is otherwise the same as embodiment 1.
Example 3
In this example, the switching circuit 8 was used instead of the relay switching circuit, and the time taken for the apparatus to monitor the same biochip sample was shortened by 0.5 minute as compared with example 1, as in example 1.
Claims (7)
1. A device for monitoring micro-array biomolecular reaction with high flux by using a light reflection difference method comprises an incident light path, a sample stage, an emergent light path and a data acquisition and processing system, wherein the incident light path comprises a laser, a polarizer, a photoelastic modulator and a phase shifter, wherein the polarizer, the photoelastic modulator and the phase shifter are sequentially arranged on the light path in front of the output light of the laser; the emergent light path comprises an analyzer and a photoelectric signal converter, the analyzer and the photoelectric signal converter are sequentially arranged in front of an emergent light beam reflected by a sample on the sample stage, the photoelectric signal converter is electrically connected to the data acquisition and processing system, and the device is characterized in that,
the incident light path also comprises a beam expanding device arranged between the phase shifter and the sample stage;
the mechanical joint is composed of two vertical plates vertically fixed on the sample table, an aluminum alloy flat plate for receiving incident light and an aluminum alloy flat plate for reflecting emergent light, wherein the two aluminum alloy flat plates and the two vertical plates are provided with arc-shaped fixing screw holes, the two aluminum alloy flat plates are obliquely fixed on the vertical plates through a bolt, the screw holes are in a quarter arc shape, and the aluminum alloy plate can be adjusted between 0 and 90 degrees;
the incident light path is arranged on the aluminum alloy flat plate for receiving incident light, and the emergent light path is arranged on the aluminum alloy flat plate for reflecting emergent light;
the photoelectric signal converter comprises a silicon photodiode array, a switching circuit, a first phase-locked amplifier and a second phase-locked amplifier, wherein the output end of the silicon photodiode array is electrically connected to the input ends of the switching circuit, the first phase-locked amplifier and the second phase-locked amplifier respectively.
2. The apparatus for high throughput monitoring of microarray biomolecular reactions using light reflectance differentials as claimed in claim 1, wherein said data acquisition and processing system comprises a BNC adapter, a data acquisition card and a data processing device; wherein,
the BNC adapter receives the output signals of the first phase-locked amplifier and the second phase-locked amplifier, performs channel division processing on the output signals of the phase-locked amplifiers, and divides the output signals of each phase-locked amplifier into two parts, namely amplitude information and phase information;
the data acquisition card acquires data output by the BNC adapter and transmits the data to the data processing device;
and the data processing device stores, analyzes and processes the data sent by the data acquisition card.
3. The device for high-throughput monitoring of the micro-array biomolecule reaction by the light reflectance difference method according to claim 1, wherein the beam expanding device is a beam expanding lens group, the beam expanding lens group comprises a collimating lens barrel and two continuous zoom beam expanding lenses respectively arranged at two ends of the collimating lens barrel, and the diameter of the cross section of the emergent beam can be adjusted by adjusting the length of the collimating lens barrel.
4. The apparatus for high throughput monitoring of microarray biomolecular reactions using light reflectance differentiation according to claim 1, wherein the sample stage is a stainless steel platform with three-dimensional manual adjustment with a sample fixing mechanism.
5. The apparatus for high throughput monitoring of reactions of biomolecules in microarrays according to claim 1, wherein said laser is a linearly polarized He-Ne laser having a wavelength of 632.8 nm.
6. The device for high throughput monitoring of the reaction of biomolecules in a microarray according to claim 1, wherein said photoelastic modulator has a modulation frequency Ω of 50 kHz.
7. The apparatus for high throughput monitoring of the reaction of biomolecules in a microarray according to claim 1, wherein said switching circuit is a relay switching circuit or a pulse switching circuit.
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| CNA2006101135764A CN101153869A (en) | 2006-09-30 | 2006-09-30 | Device for high-throughout monitoring micro-array biomolecule reaction by light reflection difference method |
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| CNA2006101135764A CN101153869A (en) | 2006-09-30 | 2006-09-30 | Device for high-throughout monitoring micro-array biomolecule reaction by light reflection difference method |
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Cited By (15)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102192879A (en) * | 2010-03-17 | 2011-09-21 | 中国科学院物理研究所 | Unmarked high throughput detecting device of biological chip by using light reflection differential method |
| CN102262052A (en) * | 2010-05-26 | 2011-11-30 | 中国科学院理化技术研究所 | Laser confocal oblique incidence ellipsometric high-flux biomolecular reaction imaging detection device |
| CN104406544A (en) * | 2014-11-20 | 2015-03-11 | 北京航空航天大学 | Detection device and method for eliminating photoelastic modulator and environment influence based on double beam difference |
| CN105424607A (en) * | 2015-12-25 | 2016-03-23 | 中国石油大学(北京) | CT device and CT method based on inclined incident light reflection difference method |
| CN105424654A (en) * | 2015-12-25 | 2016-03-23 | 中国石油大学(北京) | High-spatial-resolution light reflection difference device and high-spatial-resolution light reflection difference method used for microstructure detection |
| CN105547947A (en) * | 2015-09-23 | 2016-05-04 | 中国石油大学(北京) | Light reflection difference device and method for detecting PM2.5 |
| CN105548084A (en) * | 2015-09-23 | 2016-05-04 | 中国石油大学(北京) | Light reflection difference device and method for detecting rock stress sensitivity |
| CN105571515A (en) * | 2015-12-25 | 2016-05-11 | 中国石油大学(北京) | Method for detecting three-dimensional structure of sample by oblique-incidence reflectivity difference method |
| CN105628620A (en) * | 2015-09-23 | 2016-06-01 | 中国石油大学(北京) | Light reflex difference device and method for water flooding dynamic detection |
| CN105675542A (en) * | 2015-08-24 | 2016-06-15 | 温州大学 | Device and method for rapid identification of swill-cooked dirty oil |
| CN105717018A (en) * | 2015-09-23 | 2016-06-29 | 中国石油大学(北京) | Device and method using light reflection difference to detect rock pore structure |
| CN105717049A (en) * | 2015-09-23 | 2016-06-29 | 中国石油大学(北京) | Light reflection difference device and method for detecting oil content of oil shale |
| CN105738321A (en) * | 2015-09-23 | 2016-07-06 | 长江大学 | Light reflection difference method for detecting illegal cooking oil |
| CN105738290A (en) * | 2015-09-23 | 2016-07-06 | 长江大学 | Device and method for representing and recognizing crude oil from different producing places through light reflex difference method |
| CN105738291A (en) * | 2015-09-23 | 2016-07-06 | 长江大学 | Light reflection difference device and method for detecting anisotropies of rocks |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102192879A (en) * | 2010-03-17 | 2011-09-21 | 中国科学院物理研究所 | Unmarked high throughput detecting device of biological chip by using light reflection differential method |
| CN102192879B (en) * | 2010-03-17 | 2013-05-22 | 中国科学院物理研究所 | Unmarked high throughput detecting device of biological chip by using light reflection differential method |
| CN102262052A (en) * | 2010-05-26 | 2011-11-30 | 中国科学院理化技术研究所 | Laser confocal oblique incidence ellipsometric high-flux biomolecular reaction imaging detection device |
| CN102262052B (en) * | 2010-05-26 | 2012-12-26 | 中国科学院理化技术研究所 | Laser confocal oblique incidence ellipsometric high-flux biomolecular reaction imaging detection device |
| CN104406544A (en) * | 2014-11-20 | 2015-03-11 | 北京航空航天大学 | Detection device and method for eliminating photoelastic modulator and environment influence based on double beam difference |
| CN104406544B (en) * | 2014-11-20 | 2017-04-12 | 北京航空航天大学 | Detection device and method for eliminating photoelastic modulator and environment influence based on double beam difference |
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| CN105548084A (en) * | 2015-09-23 | 2016-05-04 | 中国石油大学(北京) | Light reflection difference device and method for detecting rock stress sensitivity |
| CN105628620A (en) * | 2015-09-23 | 2016-06-01 | 中国石油大学(北京) | Light reflex difference device and method for water flooding dynamic detection |
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| CN105717049A (en) * | 2015-09-23 | 2016-06-29 | 中国石油大学(北京) | Light reflection difference device and method for detecting oil content of oil shale |
| CN105738321A (en) * | 2015-09-23 | 2016-07-06 | 长江大学 | Light reflection difference method for detecting illegal cooking oil |
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| CN105738291A (en) * | 2015-09-23 | 2016-07-06 | 长江大学 | Light reflection difference device and method for detecting anisotropies of rocks |
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Application publication date: 20080402 |