CN102072896B - Device and method for identifying protein based on catalyzed light emission on surface of nano material - Google Patents
Device and method for identifying protein based on catalyzed light emission on surface of nano material Download PDFInfo
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- CN102072896B CN102072896B CN2010105309583A CN201010530958A CN102072896B CN 102072896 B CN102072896 B CN 102072896B CN 2010105309583 A CN2010105309583 A CN 2010105309583A CN 201010530958 A CN201010530958 A CN 201010530958A CN 102072896 B CN102072896 B CN 102072896B
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Abstract
The present invention belongs to the technical field of a sensor array. A method is characterized in that protein solution is dropped onto a plurality of electric heating ceramic plates coated with different nano materials, the ceramic plates are powered up to be heated in sequence, oxidation reaction occurs between proteins and oxygen flowing in through an air inlet under effect of catalysis of the nano materials, excited-state products or intermediate-state products are produced, energy generated from an excited state to a ground state is transmitted in a form of light emission, and detection of the proteins can be completed through detection of chemiluminescence signals. The present invention also provides a device for implementation of the method, the device comprises a quartz packaging component, the quartz packaging component is provided with the air inlet, an air outlet and a sealing cover, and a photomultiplier tube is arranged on the quartz packaging component and is connected with a signal processing circuit; and the electric heating ceramic plates coated or sintered with the different nano materials on the surfaces are arranged in the component, are electrically connected in parallel and are also connected with an external direct-current power supply. The device has the advantages of simple structure, low energy consumption, low thermal background, no reagent loss, good repeatability, long service life and the like.
Description
Technical field
The invention belongs to the sensor array technology field.Be particularly related to a kind of nano-material surface-catalyzed luminescent that utilizes and discern common method of protein and sensor special array thereof.
Background technology
At present, imitation animal sense of smell or gustatory organ and the interaction response sensor array developed are listed in the application of gas detection and solution identification and have all obtained research and development in various degree.But; Being used for solution example, to discern the kind of the sensor array of biomacromolecule identification such as albumen particularly be not a lot; And the overwhelming majority is based on strong interaction between chromogenic reaction, fluorescent quenching, indicator displacement equimolecular; Signal response is difficult to reply, and causes sensor array can only do disposable use, does not meet " Electronic Nose (tongue) " Developing Trend in Technology.In addition, a lot of sensor array element have all been used colored indicator, organic fluorescent dye, the relatively poor material of macromolecule acceptor equistability, have limited the practical application of this type sensor.In recent years; We have developed the nano catalytic luminescence gas sensor that produces chemiluminescent principle based on the determinand molecule in the nano-material surface generation redox reaction with catalytic activity; Have that selectivity is good, response speed is fast, it is fast to recover, sensing material is lossless, the characteristics of long service life; And the method for utilizing luminescence imaging has made up the array of nano material can discern detection to volatile organic matters such as ethanol, trimethylamine, benzaldehydes, however can't be applied to some biomacromolecules etc. not volatile substances for example the identification of protein go in detecting.
Summary of the invention
The purpose of this invention is to provide a kind of sensor device that can identification of protein solution kind.
Device based on the nano-material surface-catalyzed luminescent identification of protein; It is characterized in that; Contain the quartz packaged assembly; The quartz packaged assembly is provided with air intake opening, gas outlet and gland bonnet, above said quartz packaged assembly, photomultiplier is set, and said photomultiplier connects signal processing circuit; Placement surface coating or sintering have six or six thermoelectric (al) sensing element arrays that above electrothermal ceramic plate constitutes of different nano materials in said quartz packaged assembly; Parallel connection on electric between the said electrothermal ceramic plate, the lead-in wire that electrothermal ceramic plate connects passes said quartz packaged assembly, connects external dc power.
Said nano material is Pt-Ba-Al, MgO, ZrO
2, γ-Al
2O
3, MgCO
3, SrCO
3, BaO, CaO, SrO and La
2O
3In any six kinds or more than.
Above said quartz packaged assembly, be positioned at the photomultiplier below and place a monochromator.
Method based on the nano-material surface-catalyzed luminescent identification of protein is characterized in that, contains following steps:
1) preparation testing protein solution, concentration is 5-50 μ g/mL;
2) nano material more than six kinds or six kinds is coated in respectively as for the electrothermal ceramic plate array surface in the quartz packaged assembly with powder or sintering processing;
3) the electric ceramic chip arrays is warming up to 500 ℃ ± 50 ℃ degree, makes the electrothermal ceramic plate activation;
4) in the quartz packaged assembly, feed reacting gas;
5) protein solution for preparing is added drop-wise to said electrothermal ceramic plate array surface; Treat that the moisture evaporation in the protein solution is dried, again the electric ceramic chip arrays switched on successively that voltage is at 65V ± 5V; Make the electrothermal ceramic plate surface be warming up to 200 ℃ ± 20 ℃; Protein under the catalysis of nano material with air generation oxidation reaction, produce the product or the intermediate state of excited state, energy sent with luminous mode when excited state was got back to ground state;
6) photomultiplier with the light amplification of sending after, optical signal transmission is handled to signal processing apparatus, accomplish the detection of protein.
Said reacting gas is pure air or N
2With O
2Mixed gas with the arbitrary proportion mixing.
Said nano material is Pt-Ba-Al, MgO, ZrO
2, γ-Al
2O
3, MgCO
3, SrCO
3, BaO, CaO, SrO and La
2O
3In any six kinds or more than.
Beneficial effect, the device and method based on the nano-material surface-catalyzed luminescent identification of protein provided by the invention can come the effective recognition different proteins, has reached its intended purposes.
Description of drawings
Fig. 1 is based on chemiluminescent apparatus structure front elevation of nano-material surface and sensor array vertical view.
Fig. 2 is the catalytic luminescence signal graph of 8 kinds of protein aqueous solutions, and six peaks of every figure represent this albumen successively at Pt-Ba-Al respectively, MgO, ZrO
2, γ-Al
2O
3, MgCO
3, SrCO
3Catalytic luminescence signal on these six kinds of nano materials.
Fig. 3 is the catalytic luminescence intensity column diagrams of 8 kinds of protein aqueous solutions on six kinds of nano materials.
Fig. 4 each albumen typicalness scoring scatter diagram for obtaining through linear discriminant analysis (LDA).
Fig. 5 is 8 kinds of protein aqueous solutions catalytic luminescence signal graphs on Pt-Ba-Al under the variable concentrations condition, the catalytic luminescence signal when on behalf of this protein concentration, six peaks of every figure be 5,10,20,30,40,50 μ g/mL respectively.
Embodiment:
Recognition Protein sensor array provided by the invention comprises the quartz packaged array component that has air intake opening, gas outlet and gland bonnet; Wherein, in the inner chamber of said quartz packaged array component, be provided with 6 array of sensor elements that above electrothermal ceramic plate constitutes that surface applied or sintering have nano material; The two ends of the electrothermal ceramic plate of each sensing unit are individually fixed on the electrode; Said each electrode passes said gland bonnet with lead-in wire, is connected on the circuit of darkroom outer control heating sequence and heating-up temperature.
In this sensor, the various nano materials that protein is had a catalytic luminescence characteristic all are applicable to this sensor, like Pt-Ba-Al, and MgO, ZrO
2, γ-Al
2O
3, MgCO
3, SrCO
3Deng; The various common higher electrothermal ceramic plates of heating temp all are applicable to this sensor.This sensor also comprises the optical signal receiver of being located at said quartz packaged assembly top, and this optical signal receiver can be made up of monochromator and photomultiplier.Maximum voltage and maximum current between the power of electrothermal ceramic plate and size, two electrodes all can be chosen according to actual needs.
The method of Recognition Protein provided by the invention; Be that protein aqueous solution (5-50 μ g/ml) is added drop-wise on each array element (being coated with the electrothermal ceramic plate of different nano materials); Treat in the solution that moisture evaporation does, again with the heating plate energising, the air generation oxidation reaction that protein is just come under the catalysis of nano material and from air intake opening; Produce the product or the intermediate state of excited state; Energy sent with luminous mode when excited state was got back to ground state, through detecting this chemiluminescence signal, accomplished the detection of protein.Simultaneously, through 6 array elements of heated in sequence, the mode of acquisition order luminous signal; We can obtain the catalytic luminescence response signal mode chart of protein on array; Utilize again computing machine carry out linear discriminant analysis (Linear Discriminant Analysis, LDA) or principal component analysis (PCA) (Principal Component Analysis, PCA); Obtain the differentiation classification results of each protein aqueous solution, realize that the identification of multiple proteins detects.
This method is to utilize the protein in the nano material catalysis WS to produce emitting characteristics, presses the array recognized patterns, and each protein aqueous solution is differentiated.The temperature of reaction of catalytic reaction is provided by the electrothermal ceramic plate of heating, concrete temperature of reaction be albumen under nano material catalysis and the temperature of air generation oxidation reaction, as 150-300 ℃, preferred 205 ℃.The flow velocity of bubbling air is 75-375mL/min, preferred 200mL/min, and air also can be by N
2With O
2The combination gas that mixes with arbitrary proportion substitutes.
The present invention utilizes nano-material surface protein to be had the characteristics of catalytic luminescence; And different materials is to the catalytic luminescence intensity of albumen of the same race difference to some extent; The catalytic luminescence intensity of different albumen on same material is also different; Can form " finger print information " foundation, and then make up the nano material array, realize the identification of different proteins is detected as identification of protein.This sensor array is simple in structure, also has low, the no reagent loss of energy consumption and hot background, good reproducibility, long characteristics of life-span simultaneously, can be used as " electronic tongues " protein aqueous solution is discerned.
Below in conjunction with description of drawings embodiment of the present invention.
Fig. 1 is the chemiluminescent sensor array structural representation of nano-material surface provided by the invention.Being constructed as follows of this sensor array: six groups of electrothermal ceramic plates 5 and (design of circuit board makes six groups of electrothermal ceramic plates parallel connections on the sensing element fixing circuit board that its surface applied nano material 6 constitutes; And be equipped with voltage control system respectively); Two lead-in wires 7,8 pass gland bonnet 3, place the quartz packaged assembly 4 that has air intake opening 1 and gas outlet 2.Wherein, the circuit connection makes six groups of electrothermal ceramic plates to heat successively.In addition, this device also comprises the optical signal receiver on quartz packaged assembly top: monochromator 9 and photomultiplier 10.
The electrothermal ceramic plate size of used sensor is 10 * 10mm among the following embodiment
2/ sheet also can be transferred the potsherd area greatly according to actual needs, but surpass 300mm
2/ sheet is applied to voltage between the every electrothermal ceramic plate in proper range.
The concrete testing process of the identification of protein WS provided by the invention is following:
With nano material 6 apply according to conventional method with the form of powder or film or sintering on the surface of electrothermal ceramic plate 5, be heated to 500 ℃ or more than, keep more than the temperature 10min, to eliminate the interference of adsorbate.On the nano-material surface of each sensing unit, drip 10 μ L (also can be 5-20 μ L) protein aqueous solution earlier; Treat in the solution that moisture evaporation does, again with the heating plate energising, the air generation oxidation reaction that protein is just come under the catalysis of nano material and from air intake opening 1; Produce the product or the intermediate state of excited state; Energy sent with luminous mode when excited state was got back to ground state, amplified the back computer through photomultiplier and write down this chemiluminescence signal automatically, accomplished the detection of protein.To the range protein WS that will test with identical method repeated test 5-7 time; At last every kind of protein aqueous solution is delivered to data handling system at the catalytic luminescence mode chart of nano material array surface; Through LDA or PCA calculation process, successfully protein is distinguished and identification.
Below in conjunction with specific embodiment the present invention is described further, but the present invention is not limited to following examples.
With six kinds of nano material Pt-Ba-Al, MgO, ZrO
2, γ-Al
2O
3, MgCO
3, SrCO
3Form with powder is coated in six heating ceramic sheet surfaces respectively, is warming up to 500 ℃, keeps 10min, to eliminate the interference of adsorbate.Circulation gas gas is pure air (being taken place by pneumatic pump), and flow velocity is about 100mL/min.Get 10 μ L protein aqueous solutions,, be added drop-wise to the surface of each nano material like 10 μ L bovine serum albumin(BSA) (BSA) WS (concentration can at 5-50 μ g/mL); Treat that moisture evaporation is done in the solution, again six heating plates switched on successively that voltage is at 65V ± 5V; This moment, nano-material surface can produce the high temperature about 205 ℃; Protein just under the catalysis of nano material and from the air generation oxidation reaction that air intake opening is come in, produces the product or the intermediate state of excited state, and energy sent with luminous mode when excited state was got back to ground state; Amplify the back computer through photomultiplier and write down this chemiluminescence signal automatically, accomplish the detection of protein.(obtain 6 kinds different?)
With six kinds of nano material Pt-Ba-Al, MgO, ZrO
2, γ-Al
2O
3, MgCO
3, SrCO
3Form with powder is coated in six heating ceramic sheet surfaces respectively, is warming up to 500 ℃, keeps 10min, to eliminate the interference of adsorbate.Circulation gas gas is pure air (being taken place by pneumatic pump), and flow velocity is about 100mL/min.The preparation bovine serum albumin(BSA), human serum albumins, oralbumin, lysozyme, ribonuclease, trypsase, pepsin, the WS of these eight kinds of albumen of papain, concentration all is controlled at 10 μ g/mL.The WS to each protein all is added drop-wise to each array element surface in advance, treats that moisture evaporation is done in the solution, heating plate is switched on again; 65V ± 5V; Protein just under the catalysis of nano material and from the air generation oxidation reaction that air intake opening is come in, produces the product or the intermediate state of excited state, and energy sent with luminous mode when excited state was got back to ground state; Through detecting this chemiluminescence signal, accomplish the detection of protein.Simultaneously, through 6 array elements of heated in sequence, the luminous signal that every kind of protein is produced on 6 kinds of nano materials carries out acquisition order, and we can obtain the catalytic luminescence response signal mode chart (Fig. 2) of protein on array.These 8 kinds of protein are all carried out aforesaid operations and test.
With the experiment repeated several times (being generally 5-7 time) among the embodiment 2; Obtain the fingerprint spectrogram of 8 kinds of protein; We can be clear that very that different materials is to the catalytic luminescence intensity of albumen of the same race difference to some extent from Fig. 3; The catalytic luminescence intensity of different albumen on same material is also different, and the difference between each albumen comes into plain view.The catalytic luminescence intensity raw data input computer that again each protein aqueous solution is listed at sensor array; Carrying out LDA with SPSS v16.0 (also can use other statistical softwares such as SYSTAT) analyzes; Obtain each albumen typicalness scoring scatter diagram like Fig. 4; Figure can find out for albumen of the same race thus, and five repeated experiments obtain 5 typicalness scoring points and all gather in the less area of space, and the scoring of the typicalness between different albumen space of points distance relatively separately; Promptly be distributed in eight zones, space, accurately accomplished the identification of these eight kinds of protein aqueous solutions for our 8 kinds of albumen typicalness scoring points of test.
Embodiment 4:
The preparation bovine serum albumin(BSA), human serum albumins, oralbumin; Lysozyme, ribonuclease, trypsase; Pepsin, the WS of these eight kinds of albumen of papain is all prepared the WS that six routine concentration branches are respectively 5,10,20,30,40,50 μ g/mL for every kind of albumen.Single the kind on the nano material of Pt-Ba-Al these 8 kinds of albumen are measured enriched sample from low concentration sample respectively according to embodiment 2 described methods then, obtain signal results like Fig. 5.Can find out that from this result the catalytic luminescence intensity when concentration changes from low to high for these eight kinds of albumen on nano material also changes from low to high, explain that this achievement can distinguish variable concentrations albumen.
Claims (3)
1. based on the device of nano-material surface-catalyzed luminescent identification of protein; It is characterized in that; Contain the quartz packaged assembly; The quartz packaged assembly is provided with air intake opening, gas outlet and gland bonnet, above said quartz packaged assembly, photomultiplier is set, and said photomultiplier connects signal processing circuit; Placement surface coating or sintering have six or six thermoelectric (al) sensing element arrays that above electrothermal ceramic plate constitutes of different nano materials in said quartz packaged assembly; Parallel connection on electric between the said electrothermal ceramic plate, the lead-in wire that electrothermal ceramic plate connects passes said quartz packaged assembly, connects external dc power; Said nano material is Pt-Ba-Al, MgO, ZrO
2, γ-Al
2O
3, MgCO
3, SrCO
3, BaO, CaO, SrO and La
2O
3In any six kinds or more than.
2. the device based on the nano-material surface-catalyzed luminescent identification of protein as claimed in claim 1 is characterized in that, above said quartz packaged assembly, is positioned at the photomultiplier below and places a monochromator.
3. based on the method for nano-material surface-catalyzed luminescent identification of protein, it is characterized in that, contain following steps:
1) preparation testing protein solution, concentration is 5-50 μ g/mL;
2) nano material more than six kinds or six kinds is coated in the electrothermal ceramic plate array surface that places in the quartz packaged assembly respectively with powder or sintering processing; Said nano material is Pt-Ba-Al, MgO, ZrO
2, γ-Al
2O
3, MgCO
3, SrCO
3, BaO, CaO, SrO and La
2O
3In any six kinds or more than;
3) the electric ceramic chip arrays is warming up to 500 ℃ ± 50 ℃ degree, makes the electrothermal ceramic plate activation;
4) in the quartz packaged assembly, feed reacting gas; Said reacting gas is pure air or N
2With O
2Mixed gas with the arbitrary proportion mixing;
5) protein solution for preparing is added drop-wise to said electrothermal ceramic plate array surface; Treat that the moisture evaporation in the protein solution is dried, again the electric ceramic chip arrays switched on successively that voltage is at 65V ± 5V; Make the electrothermal ceramic plate surface be warming up to 200 ℃ ± 20 ℃; Protein under the catalysis of nano material with reacting gas generation oxidation reaction, produce the product or the intermediate state of excited state, energy sent with luminous mode when excited state was got back to ground state;
6) photomultiplier with the light amplification of sending after, optical signal transmission is handled to signal processing apparatus, accomplish the detection of protein.
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| CN102393390B (en) * | 2011-09-16 | 2013-06-05 | 广州大学 | Method for identifying organic matter by using single cataluminescence sensor |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0175577A2 (en) * | 1984-09-21 | 1986-03-26 | Jonathan L Kiel | Microchemiluminescent assay system |
| CN101793899A (en) * | 2009-02-04 | 2010-08-04 | 中国科学院电子学研究所 | Optical biosensor for detecting brain natriuretic peptide (BNP) and preparation method of reagent thereof |
| CN101858867A (en) * | 2009-04-08 | 2010-10-13 | 清华大学 | Method and sensor for nano material enrichment and in-situ chemical luminescence detection |
-
2010
- 2010-10-29 CN CN2010105309583A patent/CN102072896B/en not_active Expired - Fee Related
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0175577A2 (en) * | 1984-09-21 | 1986-03-26 | Jonathan L Kiel | Microchemiluminescent assay system |
| CN101793899A (en) * | 2009-02-04 | 2010-08-04 | 中国科学院电子学研究所 | Optical biosensor for detecting brain natriuretic peptide (BNP) and preparation method of reagent thereof |
| CN101858867A (en) * | 2009-04-08 | 2010-10-13 | 清华大学 | Method and sensor for nano material enrichment and in-situ chemical luminescence detection |
Non-Patent Citations (4)
| Title |
|---|
| Hao Kong，et al..A thermochemiluminescence array for recognition of protein subtypes and their denatured shapes.《Analyst》.2011,第136卷(第18期),3643-3648. * |
| Keith R. Millington,et al..Chemiluminescence from thermal oxidation of amino acids and proteins.《Amino Acids》.2010,第38卷1395-1405. * |
| Na Na，et al..A Catalytic Nanomaterial-Based Optical Chemo-Sensor Array.《J. AM. CHEM. SOC.》.2006,第128卷14420-14421. * |
| Yayan Wu,et al..Discrimination and Identification of Flavors with Catalytic Nanomaterial-Based Optical Chemosensor Array.《Analytical chemistry》.2009,第81卷(第3期),961-966. * |
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