CN101033483B - Sensing analytical method for substrate concentration - Google Patents
Sensing analytical method for substrate concentration Download PDFInfo
- Publication number
- CN101033483B CN101033483B CN2007100214227A CN200710021422A CN101033483B CN 101033483 B CN101033483 B CN 101033483B CN 2007100214227 A CN2007100214227 A CN 2007100214227A CN 200710021422 A CN200710021422 A CN 200710021422A CN 101033483 B CN101033483 B CN 101033483B
- Authority
- CN
- China
- Prior art keywords
- transition metal
- substrate
- concentration
- analytical method
- strawberry shape
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Images
Landscapes
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The technology of this invention is that: it mixes the transition metal nanoparticles with the amino-colloidal to achieve the strawberry-shaped colloidal complex. Then it mixes the above complexes, transition metal salts and the enzymes catalizing substrate to produce reductive product, and then adds the measured samples into mixture for reaction, in which, the final concentration of strawberry-shaped colloidal complex is 0.01-10X10<10> piece/ml, transition metal salts is 0.001-25 mu M/ml, enzyme 0.01-100 mu g/ml. The above products reduce the transition metal salt to be transition metal and deposits onto the surface of complex forming nano-shell. The formed shell is characterized to calculate the substrate concentration of sample. This invention has a higher sensitivity and lower detectionfloor.
Description
One, technical field
The invention belongs to bioanalysis and biosensor technique field, particularly a kind of sensing analytical method of concentration of substrate.
Two, background technology
Prior art: nanometer biotechnology has arrived the period of fast development, and the relevant problem of biomolecules-metal nanoparticle has obtained the concern of height in this field.Biomolecules-metal nanoparticle mixture can be used as the mark of optical pickocff, also can be used as the signal that catalytic label amplifies electricity in the bio-identification reaction, electrochemistry, trace analysis.For example, in the gold nano grain of nucleic acid functionization, gold grain interacts to produce and waits the mispairing that can be used to analyzing DNA and single base from the variation of resonance absorbing peak.Cancellation takes place with interacting as the dna structure of molecular beacon in the gold nano grain of fluorophor in binding molecule, and molecular beacon opens later fluorescence and activate, and the both can be used to analyzing DNA.The surface enhanced Raman scattering spectrum (SERS) of the gold nano grain of binding molecule can be used to amplify nucleic acid hybridization.Simultaneously, the gold nano grain of binding biomolecules can be used as the surface marker of Ag-Ab, the optical detection of DNA mixture.The catalytic performance of metallic particles makes various metals to be deposited on the nano particle by chemical process, has so just amplified the signal of biometric identification process.The gold nano grain marker that is connected with the bio-identification mixture can be grown up under catalysis, and this just can make sets up the bridging path between electrode, thereby just can detect antibody-antigenic compound, hybrid dna etc. by measuring conductivity.The metal deposition on metal nanoparticle surface can be used as mark, and the stripping voltammetry by metal refining amplifies biometric identification process.The catalysis that is connected the metal nanoparticle on the bio-identification mixture is grown up, and weight changes, and carries out responsive DNA microdetermination by quartz crystal microbalance, thereby has amplified recognition signal.The material that some oxydase energy and substrate reactions life have reductibility is as Dopamine HCL, suprarenin, H
2O
2Deng.These products can reduce AuCl under the catalysis of gold nano grain
4 -Thereby, impel gold nano grain to grow up.The variation of the optical signalling that utilizes this gold nano grain to grow up to cause can develop into the detection method (WO 2006/008742A1) that a kind of enzymic activity and substrate detect.But when existing method of directly growing up based on transition metal nanoparticles was used to analyze, its sensitivity and the detection lowest limit can not satisfy the analysis of high precision sample.The present invention is based on the transition metal nanoshell amplification enzyme catalyzed reaction of growing up, have the very high sensitivity and the lower detection lowest limit.
Three, summary of the invention
The present invention is directed to above-mentioned technical problem, a kind of sensing analytical method of concentration of substrate is provided, this method is highly sensitive, and it is low to detect bottom line.
Technical solution of the present invention is: with transition metal nanoparticles mix with amidized colloid the strawberry shape glue compound; The strawberry shape glue compound that obtains after the processing of last step is mixed with the enzyme that transition metal salt and energy catalytic substrate produce the reductibility product, add testing sample again and carry out the substrate for enzymatic activity reaction in mixed solution, wherein the final concentration of strawberry shape glue compound is 0.01-10 * 10
10Individual/ml, the final concentration of transition metal salt is 0.001-25 μ M/ml, the final concentration of enzyme is 0.01-100 μ g/ml, and the product that produces through above-mentioned reaction is reduced into transition metal with transition metal salt and is deposited on strawberry shape glue compound surface, forms the transition metal nanoshell; The transition metal nanoshell of above-mentioned formation is characterized, calculate concentration of substrate in the testing sample.
The present invention compared with prior art has following advantage:
1. the method for component is compared with the sensing assays based on nano particle in the detection of biological sample of the present invention, locals etc. are big from the migration amount of resonance peak, be 15nm based on sensing assays local of nano particle etc. to the maximum from the migration amount of resonance peak, and local of the method for component etc. is about 100nm from the migration amount maximum of resonance peak in the detection of biological sample of the present invention.
2. the method for component is compared with the sensing assays based on nano particle in the detection of biological sample of the present invention, the local of the method for component etc. can migrate near infrared region from resonance peak in the detection of biological sample of the present invention, therefore can be used for the analysis of the opaque biological sample of visible light.
Four, description of drawings
Fig. 1 is the detection reaction schema
Fig. 2 is for detecting the optical characterisation result of glucose concn, the spectrogram of the gold nanoshell of different concns glucose after by catalysis;
Fig. 3 is for detecting the optical characterisation result of glucose concn, the position at the plasmon resonance peak of the gold nanoshell of different concns glucose after by catalysis;
Fig. 4 is the typical curve that detects the electrochemical Characterization method of glucose.
Fig. 5 is the typical curve that detects the microgravity characterizing method of glucose.
Fig. 6 is the micro-characterizing method that detects tyrosine, wherein:
A:110nm/ strawberry shape glue compound;
B:121 ± 5nm/10mM tyrosine;
C:140 ± 6nm/50mM tyrosine;
D:148 ± 5nm/150mM tyrosine.
Five, embodiment
Embodiment 1 SiO
2The colloidal amination
Getting an amount of diameter is 60nm SiO
2Alcosol places a clean 100mL beaker, and (APTES Sigma) adds wherein, wraps and places on the magnetic stirring apparatus with preservative film to pipette the γ-An Jibingjisanyiyangjiguiwan of 30 μ L with pipettor.Under 80 ℃, stirred 3 hours with 600rpm.After stirring end, above-mentioned solution is placed clean centrifuge tube, with 10, the centrifugal 30mmin of 000rpm.Take out centrifuge tube bottom SiO
2Solute, the dehydrated alcohol of adding equivalent, ultra-sonic dispersion 15～20mmin.Above process repeats 5 times to remove unnecessary APTES.
The preparation of embodiment 2 gold nano grains
Utilize the method for reduction hydrochloro-auric acid (Shanghai chemical reagent one factory) to generate gold nano grain.With 1gHAuCl
4Be made into 1% the aqueous solution, 4 ℃ of refrigerations.The 40mL tri-distilled water of precooling is taken out, add the HAuCl of 0.6mL1%
4Solution adds the K that 0.2mL concentration is 0.2M again
2CO
3Solution.Under constantly stirring, add formaldehyde 0.005mL fast.The bluish voilet that is stirred to solution becomes orange red.Restir 5mmin obtains the Radioactive colloidal gold diameter generally at 10～15nm.
The preparation of embodiment 3 strawberry shape polystyrene colloid mixtures
Cut-off directly is amidized polystyrene (APTES-PS) the colloidal solution 0.07mL of 150nm, is diluted to 50mL, joins in the separating funnel of 100mL.In the 40mL of continuous stirring 10-15nm nano silver colloidal sol, dropwise add the nano silver colloidal sol of dilution, the time is about 30min.By electrostatic interaction, Nano silver grain is adsorbed onto the APTES-PS surface, forms the Ag/PS composite particles that is coated with Nano silver grain.Then, above-mentioned mixed solution is stirred 90min with 600rpm, and with 3, the centrifugal 30min of 000rpm outwells supernatant liquid gently, add tri-distilled water, ultra-sonic dispersion.Recentrifuge, ultra-sonic dispersion, triplicate.Embodiment 4 xanthine Determination on content
Get 4mL 2.4 μ M/ml hydrochloro-auric acids (the Shanghai chemical reagent one factory) aqueous solution, 0.5ml strawberry shape colloid SiO
2Mixture, concentration are 4.5 * 10
10Individual/ml and 0.5ml 4 μ g/ml XOD (Sigma) solution mixings add rapidly in the 1mL testing sample, room temperature vibration 10 minutes.Strawberry shape colloid SiO
2Mixture becomes SiO
2Gold nanoshell, solution becomes light bluely by colourless, by colorimetric method for determining xanthine content in the calculation sample as a result, sees Fig. 1.
The calculating of glucose concn is that glucose gold nanoshell plasmon resonance spike after catalysis of mensuration series amount earlier is long, the typical curve equation of match.Bring the plasmon resonance spike length of sample gained into the typical curve equation again, promptly get glucose concn in the sample.
The typical curve equation of Fig. 1 match: y=1.4951 * 10
6X+536.9682;
Bring the plasmon resonance spike length of testing sample gained into concentration that above-mentioned equation promptly gets glucose in the sample.
Fig. 2 and Fig. 3 detect the electrochemistry of glucose and the typical curve of microgravity characterizing method.Method of calculation are the same.
Embodiment 5 tyrosine Determination on content
Get 1mL 5.4 μ M/ml hydrochloro-auric acids (the Shanghai chemical reagent one factory) aqueous solution, 1ml strawberry shape colloid SiO
2Mixture (1 * 1010/ml) and 0.5ml 4 μ g/ml tyrosine oxidase (Sigma) solution mixings, add rapidly in the 0.5ml testing sample, room temperature vibration 10 minutes.Strawberry shape colloidal sio2 mixture becomes the SiO2 gold nanoshell, solution by colourless become light blue, by tyrosine content (Fig. 4) in the microscopy measurement result calculation sample.Method of calculation are with embodiment 4.
Embodiment 6 L-glutamic acid Determination on content
Get 2mL 0.01mM Silver Nitrate (the Nanjing chemical reagent one factory) aqueous solution, 1ml strawberry shape colloid PS mixture (10 * 10
10Individual/as ml) and 0.5ml 60 μ g/ml glutamate dehydrogenase (Sigma) solution mixings, to add rapidly in the testing sample, room temperature vibration 30 minutes.Strawberry shape colloid PS mixture forms PS silver nanoshell, solution by colourless become light blue, this has indicated the existence of glutamate dehydrogenase and catalysis L-glutamic acid.L-glutamic acid cubage method is with embodiment 4.
Claims (3)
1. the sensing analytical method of a concentration of substrate is characterized in that step is:
A. transition metal nanoparticles is mixed with amidized colloid the strawberry shape glue compound;
B. will go up the strawberry shape glue compound, the transition metal salt that obtain after the step processing mixes with the enzyme that the energy catalytic substrate produces the reductibility product, add testing sample again and carry out the substrate for enzymatic activity reaction in mixed solution, wherein the final concentration of strawberry shape glue compound is 0.01-10 * 10
10Individual/ml, the final concentration of transition metal salt is 0.001-25 μ M/ml, the final concentration of enzyme is 0.01-100 μ g/ml, and the product that produces through above-mentioned reaction is reduced into transition metal with transition metal salt and is deposited on strawberry shape glue compound surface, forms the transition metal nanoshell;
C. the transition metal nanoshell with above-mentioned formation characterizes, and calculates concentration of substrate in the testing sample.
2. the sensing analytical method of a kind of concentration of substrate as claimed in claim 1 is characterized in that described amidized colloidal particle diameter is 150nm.
3. the sensing analytical method of a kind of concentration of substrate as claimed in claim 1 is characterized in that described transition metal particles diameter is 10-15nm.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2007100214227A CN101033483B (en) | 2007-04-12 | 2007-04-12 | Sensing analytical method for substrate concentration |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2007100214227A CN101033483B (en) | 2007-04-12 | 2007-04-12 | Sensing analytical method for substrate concentration |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN101033483A CN101033483A (en) | 2007-09-12 |
| CN101033483B true CN101033483B (en) | 2011-09-07 |
Family
ID=38730224
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2007100214227A Active CN101033483B (en) | 2007-04-12 | 2007-04-12 | Sensing analytical method for substrate concentration |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN101033483B (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106841162B (en) * | 2017-01-19 | 2020-01-07 | 东南大学 | Method for detecting oxidation reduction level of fruit or vegetable by SERS probe |
-
2007
- 2007-04-12 CN CN2007100214227A patent/CN101033483B/en active Active
Non-Patent Citations (2)
| Title |
|---|
| 谈 勇，等.金纳米壳球体的光学特性及其应用研究进展.《科学通报》.2005,第50卷(第6期),第505－511页. * |
| 谈 勇,等.金纳米壳球体的制备及其潜在的生物学应用.《化学学报》.2005,第63卷929-933. * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN101033483A (en) | 2007-09-12 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN110658177B (en) | Phenol recognition SERS probe, preparation and application thereof, and SERS-based general ultrasensitive immunoassay method | |
| Liang et al. | Metal-enhanced ratiometric fluorescence/naked eye bimodal biosensor for lead ions analysis with bifunctional nanocomposite probes | |
| Zhao et al. | Ag/CdO NP-engineered magnetic electrochemical aptasensor for prostatic specific antigen detection | |
| Zeng et al. | Magnetic bead-based photoelectrochemical immunoassay for sensitive detection of carcinoembryonic antigen using hollow cadmium sulfide | |
| CN109612977B (en) | Enzyme-free signal amplification biomarker detection method based on surface enhanced Raman spectroscopy | |
| CN109813785B (en) | Acetamiprid electrochemical aptamer sensor based on hollow porous nanosphere double-amplification signal | |
| Haider et al. | Gold nanoparticle decorated single walled carbon nanotube nanocomposite with synergistic peroxidase like activity for d-alanine detection | |
| Li-Na et al. | Synthesis and applications of gold nanoparticle probes | |
| Li et al. | A surface enhanced Raman scattering quantitative analytical platform for detection of trace Cu coupled the catalytic reaction and gold nanoparticle aggregation with label-free Victoria blue B molecular probe | |
| CN108918853B (en) | Pd @ Ag @ CeO2Preparation method and application of labeled immunosensor | |
| Liang et al. | Immunocontrolling graphene oxide catalytic nanogold reaction and its application to SERS quantitative analysis | |
| ZHANG et al. | Single particle impact electrochemistry: analyses of nanoparticles and biomolecules | |
| CN106124588B (en) | A kind of preparation method of the electrochemistry nonyl phenol sensor based on titania-doped/molybdenum disulfide composite material | |
| CN107356583A (en) | One kind tetraphenylboron sodium part regulation and control Nano Silver catalytically active surface enhancing Raman spectroscopy NH4+Method | |
| Alijanianzadeh et al. | Detection of methamphetamine using aptamer-based biosensor chip and cyclic voltammetry technique | |
| CN112525971B (en) | Method for photoelectrochemical detection of chloramphenicol based on bismuth tungstate | |
| CN107764763B (en) | Iodine ion signal enhanced hydrogen peroxide colorimetric detection method | |
| Jiang et al. | Inhibition to dual enzyme-like activities of Ag/CeO2 nanozymes for the detection of thiourea | |
| CN110441535A (en) | A kind of preparation method of the electrochemical immunosensor based on Pd NCs functionalization CuInOS detection Procalcitonin | |
| CN101033483B (en) | Sensing analytical method for substrate concentration | |
| CN101201318B (en) | Detection of trace amount gold nanometer particle by HAuCl4 spectrophotometric method | |
| CN110596065B (en) | Acid phosphatase detection method based on cysteamine-N-acetyl-L-cysteine-gold nanocluster fluorescent material | |
| Li et al. | Selective and accurate detection of nitrate in aquaculture water with surface-enhanced raman scattering (SERS) using gold nanoparticles decorated with β-cyclodextrins | |
| CN113777088B (en) | Fluorescent detection method of acetylcholinesterase based on carbon dots | |
| Yang et al. | One-step synthesis of triethanolamine-capped Pt nanoparticle for colorimetric and electrochemiluminescent immunoassay of SARS-CoV spike proteins |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| ASS | Succession or assignment of patent right |
Owner name: SOWTHEAST UNIV. Effective date: 20140820 Owner name: NANTONG HAILITE RUBBER + PLASTIC MACHINE CO., LTD. Free format text: FORMER OWNER: SOWTHEAST UNIV. Effective date: 20140820 |
|
| C41 | Transfer of patent application or patent right or utility model | ||
| COR | Change of bibliographic data |
Free format text: CORRECT: ADDRESS; FROM: 210096 NANJING, JIANGSU PROVINCE TO: 226623 NANTONG, JIANGSU PROVINCE |
|
| TR01 | Transfer of patent right |
Effective date of registration: 20140820 Address after: 226623, No. 2, C District, Chengbei Industrial Zone, Haian County, Jiangsu City, Nantong province (Haian County Industrial Zone) Patentee after: Nantong Haylett Rubber and Plastic Machinery Co., Ltd. Patentee after: Southeast University Address before: 210096 Jiangsu city Nanjing Province four pailou No. 2 Patentee before: Southeast University |