CN103630598A - Electrochemical detection method of cocaine based on rolling circle amplification and supramolecular aptamer - Google Patents
Electrochemical detection method of cocaine based on rolling circle amplification and supramolecular aptamer Download PDFInfo
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Abstract
The invention discloses an electrochemical detection method of cocaine based on rolling circle amplification and a supramolecular aptamer. The method comprises the steps of dropwise adding an aptamer segment Co3S onto a cleaned gold electrode, taking out for sealing after overnighting in a refrigerator, dropwise adding an aptamer segment Co3B and a sample to be detected on the gold electrode, after reaction at 37 DEG C, dropwise adding a streptavidin solution for incubation, then adding a cyclized DNA (deoxyribonucleic acid) solution for incubation, performing rolling circle amplification, adding a detection probe for hybridization and performing DPV (differential pulse voltammetry) signal detection. The method utilizes the rolling circle amplification and the supramolecular aptamer for electrochemical detection of the cocaine for the first time, greatly improves sensitivity and specificity of cocaine detection, and allows a linear detection range to reach 10-500nM. The streptavidin solution and cyclized DNA in the same volume and at the same concentration are utilized, an identification element and an amplifier element are separated, a blank signal is decreased by a bridge connection effect of streptavidin is only utilized, and the sensitivity and the linear range of the detection are greatly improved and extended.
Description
Technical field
The present invention relates to a kind of detection method of cocaine, relate in particular to a kind ofly based on rolling circle amplification and the fit method of cocaine being carried out to Electrochemical Detection of supermolecule, belong to field of biological detection.
Background technology
Cocaine (cocaine) is main alkaloid contained in coca leaf, has another name called cocaine.In coca leaf, there are a lot of alkaloids, mainly contain the materials such as cocaine, ecgonine (ecgonine), benzoylecgonine, cinnamic acid cocaine and N-formaldehyde cocaine, central nervous system is had to serious toxic action.The method at present cocaine being detected has vapor-phase chromatography (GC), high performance liquid chromatography (HPLC), gas chromatography-mass spectrography (GC-MS), thin-layered chromatography (TLC), Electronic Nose, electron capture, micro-Raman technology (MRS, micro raman spectroscopy), X-ray detection technology and thermal neutron tomoscan etc.The detection sensitivity of these technology is high, but detecting step is loaded down with trivial details, apparatus expensive, and test procedure is complicated, consuming time, and cost is higher, needs the professional technique talent, is not suitable for on-site quick screening and uses.For the actual demand of drugs fighting work, need to develop highly sensitive, selectivity good, portable, energy consumption is low and easy-operating intelligent drugs fast detecting product, to reach sensitive and testing goal fast.To the major technique of illicit drugs inspection in the quick field screening of drug addict and liquid, be various based on immunoreactive product both at home and abroad at present, mainly contain EIA enzyme immunoassay and immune colloidal gold technique.The former relates to enzyme reaction, and detecting step is loaded down with trivial details, and reagent stability is poor, conventionally can not preserve for a long time.The latter is easy, and fast, but False Rate is high, and sensitivity is lower, can only carry out qualitative test, is difficult to carry out quality control, even same series-produced reagent, the also very difficult homogeneity that guarantees each reagent.Therefore, generally can only, for qualitative test, can not carry out quantitative test.
Rolling circle amplification technology (RCA) is a kind of method of the single-stranded cyclic DNA that directly increases under constant temperature.The signal that can not only realize target sequence amplifies, and increases and realize under isothermal, and reaction conditions is gentle, and its sensitivity can reach the nucleic acid molecules of 1 copy.The advantage of RCA technology is: high sensitivity, high specific, simple, easy to operate, diversity, high flux.
Aptamers (Aptamer) is from 10
15the DNA of random series or RNA storehouse in system molecular evolution method (being called for short SELEX) by index concentration screen, can be combined with target molecules specifically, thereby identify target molecules specifically.Aptamers can be for comprising the detection of the various biomolecule of protein, little molecule, heavy metal ion, cell and virus etc.Compare with traditional antibody, aptamers has significant superiority in many aspects.Aptamers stability is high, cheap, different batches product quality is identical, and has and the quite even better selectivity of antibody and compatibility.Utilize at present aptamers to carry out the method for cocaine detection, be mainly to use one section of aptamers, or use one section of aptamers that is separated into two sections is carried out to the detection of cocaine.In concrete detection method, be mainly to utilize fluorescent quenching, enzymatic color reaction, galvanochemistry or nm of gold color reaction to carry out the detection of cocaine.The detection range of linearity of these methods is in the scope of 0.1 μ M-500 μ M.
The detection method of cocaine has following several: a kind of is the method (B.R.Baker that uses one section of aptamers, et.al, J.Am.Chem.Soc.2006,128,3138-3139), in the method, a terminal modified sulfydryl of aptamers, by Au-S key, be fixed on gold electrode surfaces, the other end is modified oxidizing reducing group methylene blue (MB).In the situation that cocaine exists, cocaine and its aptamers interact, and cause conformational change, cause MB to approach electrode surface, thereby cause the increase of electrode surface electric current, realize the quantitative detection of cocaine, and the detection sensitivity of the method is 10 μ M.The second be use two-part aptamers probe (J.Am.Chem.Soc.Vol.131, No.21,2009,6944-6945.), one section of probe is assembled on electrode by sulfydryl, and by another segment mark MB.In the situation that cocaine exists, the probe that MB modifies makes MB near electrode surface by cocaine and the probe formation supramolecular complex on electrode surface, thereby cause the increase of electrode surface electric current, the sensitivity of detection has been improved to an order of magnitude (1 μ M).Patented claim CN102650612A has proposed to utilize syllogic aptamers probe to carry out Electrochemical Detection to cocaine, and it is divided into three sections by aptamers, and sensitivity is at 0.1 μ M.Patent CN 101948907B has proposed to utilize the syllogic aptamers probe of unmodified to carry out the cocaine nm of gold method for detecting color of homogeneous phase, the method is by the cocaine nm of gold color detection based on a section or two-part aptamers, and sensitivity is brought up to 0.5 μ M by 2.5 μ M.This patented invention has confirmed one section of aptamers probe to be divided into after three sections first, still can form and have double-stranded DNA structure with cocaine self assembly.The above adopts the syllogic aptamers probe of unmodified to carry out the method for cocaine color detection, and adopt the method that a section or two sections of aptamers are carried out cocaine Electrochemical Detection to exist following deficiency: to there is significant limitation, detection sensitivity not high (0.5 μ M～10 μ M level), standard deviation is large, and quantitative test error is large.
Summary of the invention
For above problem, it is a kind of highly sensitive based on rolling circle amplification with supermolecule is fit that cocaine is carried out to electrochemical detection method that technical matters to be solved by this invention is to provide.
For achieving the above object, the technical solution used in the present invention is: a kind of based on rolling circle amplification and the fit method to cocaine Electrochemical Detection of supermolecule, comprise the following steps:
1) by naked gold electrode polishing, washing, then immerses 10～15min in Piranha solution, then takes out and clean, be dried;
2) the fit fragment Co3S of the marking sulfhydryl with the preparation of TE damping fluid is dripped on above-mentioned gold electrode, be placed in 4 ℃ of refrigerator overnight;
3) gold electrode in step 2 is taken out, rinse, with 6-sulfydryl-1-hexanol sealing 1～1.5h, again rinse gold electrode, then with salmon sperm dna and 2%BSA mixed solution sealing gold electrode 2～3h;
4) gold electrode after sealing in step 3 is taken out and rinsed, on gold electrode, drip biotin labeled fit fragment Co3B and the testing sample solution of PB damping fluid preparation, then at 37 ℃, react 40～50min;
5) gold electrode in step 4 is taken out and rinsed, add the streptavidin solution of PB damping fluid preparation, 37 ℃ of incubation 30～45min, take out and rinse;
6) on the gold electrode in step 5, add the cyclized DNA solution of PB damping fluid preparation, 37 ℃ of incubation 30～45min, take out and rinse;
7) on the gold electrode in step 6, add the rolling circle amplification reactant liquor containing dNTP, phi29DNA polymerase, 37 ℃ of rolling circle amplification 1～1.5h, take out and rinse;
8) on the gold electrode in step 7, add the detector probe with the biotin modification of 2 * SSC hybridization solution preparation, 37 ℃ of hybridization 1～1.5h, with DEA buffer solution for cleaning electrode;
9) on the gold electrode in step 8, add the ST-AP with 1.25 μ g/mL of the DEA damping fluid preparation that contains 0.8%BSA, 37 ℃ of reaction 30～45min, with DEA damping fluid, rinse, in the 0.75mg/mL α-NP substrate solution with the preparation of DEA damping fluid, carry out Differential Pulse Voltammetry DPV input, according to typical curve, obtain the concentration of cocaine in sample to be checked.
In described step 1, gold electrode polishing is the alumina powder of 0.05 μ m by granularity, and described Piranha solution is dense H
2sO
4: H
2o
2volume ratio is the solution of 3:1.
The fit fragment Co3S sequence of described marking sulfhydryl is: 5 '-GGGAGTCAAGAACGAAAAAAAA-thiol-(CH
2)
3biotin labeled fit fragment Co3B sequence is: 5 '-TTCGTTCTTCAATGAAGTGGGACGACAAAAAAA-biotin, detector probe sequence is: 5 '-Biotin-AAAAAAGCGCAGAATGGT, while making cyclized DNA, primed DNA sequence used is: 5 '-Biotin-AAAAAAAAAAAACAGGGCTGGGCATAGAAGTCAGGGCAGAGA, treats that cyclisation template DNA sequence is: 5 '-P-TATGCCCAGCCCTGTAAGATGAAGATAGCGCAGAATGGTCGGATTCTCAACTCG TATCTGCCCTGACTTC.
Described step 3 to the flushing of gold electrode in 7 completes in accordance with the following steps: first with rinsing three times containing 0.005% polysorbas20 Tris-HCl damping fluid, and then with Tris-HCl damping fluid flushing three times; Tris-HCl damping fluid is for containing 20mM Tris, 0.1M sodium chloride, and 5mM magnesium chloride, pH 7.4.
The salmon sperm dna that salmon sperm dna in described step 3 and 2%BSA mixed solution are 10mg/ml by concentration and 2%BSA volume ratio are 1:80 mixed preparing.
The preparation method of the testing sample solution in described step 4 is: urine is centrifugal, get supernatant, then with the 1:3 dilution by volume of PB damping fluid; PB damping fluid in described step 4,5,6 is for containing 5mM potassium chloride, 5mM magnesium chloride, and 40 μ M TCEP, 0.1M phosphate, pH 7.4.
In rolling circle amplification reactant liquor in described step 7, contain 33mM Tris, 10mM magnesium chloride, 66mM potassium chloride, 1mM dithiothreitol (DTT), 0.1% Tween-20, pH 7.9.
Hybridization solution in described step 8 is 2 * SSC damping fluid, contains 0.3M sodium chloride, 0.03M trisodium citrate, and pH 7.4; DEA damping fluid is for containing 0.1M diethanolamine, 1mM magnesium chloride, and 100mM potassium chloride, pH 9.6.
The concentration of the streptavidin solution of preparing with PB damping fluid in described step 5 is identical with the cyclized DNA concentration in step 6, and the amount adding is also identical.Use streptavidin solution and the cyclized DNA of same volume and same concentrations, recognition component and amplifier element are separated, only utilize the bridge joint effect of streptavidin, reduced blank signal, improved the sensitivity and linear measurement range detecting.
The range of linearity that the method cocaine detects is 10～500nM, and sensitivity is 9.26nM, and the sensitivity that detects cocaine than existing biology sensor improves 1 order of magnitude, is conducive to trace or trace drugs fast detecting.
Beneficial effect: the present invention utilizes the fit and rolling circle amplification of supermolecule to carry out Electrochemical Detection to cocaine first, sensitivity and specificity that the fit sensor of cocaine detects have greatly been improved, make the range of linearity detecting reach 10～500nM, the sensitivity of detection is 9.26nM.Use streptavidin solution and the cyclized DNA of same volume and same concentrations, recognition component and amplifier element are separated, only utilize the bridge joint effect of streptavidin, reduced blank signal, improved the sensitivity and linear measurement range detecting.
Figure of description:
Fig. 1 is detection principle schematic of the present invention;
Fig. 2 is cocaine concentration of the present invention and current signal linear graph of a relation;
Fig. 3 is the specificity investigation figure of fit sensor of the present invention, and wherein a is blank, and b is morphine, and c is crystal methamphetamine, and d is ketamine, and e is cocaine.
Embodiment
Below in conjunction with drawings and Examples, the invention will be further described:
ST-AP in the present invention is Streptavidin-alkaline Phosphatase, and Chinese is Avidin mark alkaline phosphatase, purchased from Sigma company (U.S.).
α-NP in the present invention is α-Naphthyl Phosphate, and Chinese is 1-naphthols sodium phosphate, purchased from Sigma company (U.S.).
TCEP in the present invention is Tris (2-carboxyethyl) phosphine hydrochloride, and Chinese is the acid of three (2-carboxyethyl) microcosmic salt, purchased from Sigma company (U.S.).
BSA in the present invention is bovine serum albumin, and Chinese is bSA, purchased from Sigma company (U.S.).
Embodiment mono-, prepare cyclized DNA
The primed DNA of the biotin modification for the treatment of cyclisation template DNA and 100nM of 100nM 5 ' end phosphorylation is mixed in to 100 μ L and contains 6.6mM magnesium chloride, 10mM dithiothreitol (DTT), 1mM ATP, in the damping fluid of 66mMTris, pH 7.6, then the T4DNA ligase that adds 0.2U, 37 ℃ of coupled reaction 60min, 65 ℃ of 10min deactivation T4DNA ligases, the cyclized DNA obtaining saves backup at-20 ℃.The used cyclisation template DNA sequence for the treatment of is:
5 '-P-TATGCCCAGCCCTGTAAGATGAAGATAGCGCAGAATGGTCGGATTCTCAACTCG TATCTGCCCTGACTTC, primed DNA sequence is: 5 '-Biotin-AAAAAAAAAAAACAGGGCTGGGCATAGAAGTCAGGGCAGA.
Embodiment bis-, based on rolling circle amplification and the fit method to cocaine Electrochemical Detection of supermolecule, according to following steps, carry out:
1) 0.05 μ m alumina powder polishing for naked gold electrode, supersound washing in water, immerses Piranha solution (dense H
2sO
4: H
2o
2volume ratio is 3:1) 10～15min, washed with de-ionized water gold electrode, natural drying at room temperature;
2) the fit fragment Co3S of the 50nM marking sulfhydryl of the TE damping fluid of 10 μ L (pH 8.0,10mMTris, 1mMEDTA) preparation is dripped on the gold electrode in step 1, be placed in 4 ℃ of refrigerator overnight; The fit fragment Co3S sequence of described marking sulfhydryl is:
5’-GGGAGTCAAGAACGAAAAAAAA-thiol-(CH
2)
3；
3) with the gold electrode in cleaning buffer solution rinsing step 2, the 6-sulfydryl-1-hexanol sealing 1～1.5h with 10 μ L1mM, rinses gold electrode with cleaning buffer solution, then with 10 μ L salmon sperm dnas and 2%BSA mixed solution sealing gold electrode 2～3h; The salmon sperm dna that described salmon sperm dna and 2%BSA mixed solution are 10mg/ml by concentration and 2%BSA volume ratio are 1:80 mixed preparing.
4) with the gold electrode in cleaning buffer solution rinsing step 3, the biotin labeled fit fragment Co3B of 60nM and the 5 μ L testing sample solutions that on gold electrode, drip 5 μ L react 40～50min simultaneously at 37 ℃; Described biotin labeled fit fragment Co3B sequence is: 5 '-TTCGTTCTTCAATGAAGTGGGACGACAAAAAAA-biotin; Testing sample solution preparation method is: urine is centrifugal, get supernatant, then with the 1:3 dilution by volume of PB damping fluid; PB damping fluid in described step 4,5,6 is for containing 5mM potassium chloride, 5mM magnesium chloride, and the 0.1M phosphate of 40 μ M TCEP, pH 7.4.
5) with the gold electrode in cleaning buffer solution rinsing step 4, add the streptavidin solution with the preparation of PB damping fluid of 10 μ L15nM, 37 ℃ of incubation 30min, cleaning buffer solution rinses;
6) on the gold electrode in step 5, add 10 μ L containing the PB damping fluid of 15nM cyclized DNA, 37 ℃ of incubation 30min, rinse with cleaning buffer solution;
7) on the gold electrode in step 6, add 10 μ L containing the rolling circle amplification reactant liquor of 0.5mM dNTP, 0.4U phi29 archaeal dna polymerase, 37 ℃ of rolling circle amplification 1h, rinse gold electrode with cleaning buffer solution; In described rolling circle amplification reactant liquor, contain 33mM Tris, 10mM magnesium chloride, 66mM potassium chloride, 1mM dithiothreitol (DTT), 0.1% Tween-20, pH 7.9.
8) on the gold electrode in step 7, add 37 ℃ of hybridization 1h of detector probe of 1 μ M biotin modification of the use 2 * SSC hybridization solution preparation of 10 μ L, with DEA buffer solution for cleaning gold electrode; Described detector probe sequence is: 5 '-Biotin-AAAAAAGCGCAGAATGGT; Described hybridization solution is 2 * SSC damping fluid, contains 0.3M sodium chloride, 0.03M trisodium citrate, and pH 7.4; DEA damping fluid is for containing 0.1M diethanolamine, 1mM magnesium chloride, and 100mM potassium chloride, pH 9.6.
9) on the gold electrode in step 8, add the ST-AP of 1.25 μ g/mL of the DEA damping fluid preparation that contains 0.8%BSA for 10 μ L, 37 ℃ of reaction 30min, first with the DEA damping fluid that contains 0.005% polysorbas20, rinse three times, with the DEA damping fluid that does not contain polysorbas20, rinse three times again, in the α-NP substrate solution of the 0.75mg/mL with the preparation of DEA damping fluid, carry out DPV input.According to typical curve, obtain the concentration of cocaine in sample to be checked.
Above step 3 to the flushing of gold electrode in 7 completes in accordance with the following steps: first with rinsing three times containing 0.005% polysorbas20 Tris-HCl damping fluid, and then with Tris-HCl damping fluid flushing three times; Tris-HCl damping fluid is for containing 20mM Tris, 0.1M sodium chloride, and 5mM magnesium chloride, pH 7.4.
Electrochemical Detection DPV signal, DPV signal and cocaine concentration linear relationship chart as shown in Figure 2, calculate its linear relationship and are: y=0.02x+2.78, wherein y is current signal strength, x is cocaine concentration.According to the current signal strength value obtaining, can convert and obtain the cocaine concentration value in sample to be checked.When electric current is 4 μ A, can calculates and learn that cocaine concentration is 61nM.
Use the method to detect respectively the cocaine that morphine, crystal methamphetamine, ketamine and concentration known that blank, concentration known be 5 μ M are 500nM, obtain the fit sensor specificity investigation figure that each matter utilization the method detects, as shown in Figure 3, the concentration of morphine, crystal methamphetamine and ketamine is 10 times of cocaine, the current signal producing is but close with blank, even lower, visible the method has very high specificity to cocaine, very low to morphine, crystal methamphetamine and ketamine specificity, the method is suitable for cocaine and detects.
Claims (10)
1. based on rolling circle amplification and the fit method to cocaine Electrochemical Detection of supermolecule, comprise the following steps:
1) by naked gold electrode polishing, washing, then immerses 10～15min in Piranha solution, then takes out and clean, be dried;
2) the fit fragment Co3S of the marking sulfhydryl with the preparation of TE damping fluid is dripped on above-mentioned gold electrode, be placed in 4 ℃ of refrigerator overnight;
3) gold electrode in step 2 is taken out, rinse, with 6-sulfydryl-1-hexanol sealing 1～1.5h, again rinse gold electrode, then with salmon sperm dna and 2%BSA mixed solution sealing gold electrode 2～3h;
4) gold electrode after sealing in step 3 is taken out and rinsed, on gold electrode, drip biotin labeled fit fragment Co3B and the testing sample solution of PB damping fluid preparation, then at 37 ℃, react 40～50min;
5) gold electrode in step 4 is taken out and rinsed, add the streptavidin solution of PB damping fluid preparation, 37 ℃ of incubation 30～45min, take out and rinse;
6) on the gold electrode in step 5, add the cyclized DNA solution of PB damping fluid preparation, 37 ℃ of incubation 30～45min, take out and rinse;
7) on the gold electrode in step 6, add the rolling circle amplification reactant liquor containing dNTP, phi29DNA polymerase, 37 ℃ of rolling circle amplification 1～1.5h, take out and rinse;
8) on the gold electrode in step 7, add the detector probe with the biotin modification of 2 * SSC hybridization solution preparation, 37 ℃ of hybridization 1～1.5h, with DEA buffer solution for cleaning electrode;
9) on the gold electrode in step 8, add the ST-AP with 1.25 μ g/mL of the DEA damping fluid preparation that contains 0.8%BSA, 37 ℃ of reaction 30～45min, with DEA damping fluid, rinse, in the 0.75mg/mL α-NP substrate solution with the preparation of DEA damping fluid, carry out Differential Pulse Voltammetry DPV input, according to typical curve, obtain the concentration of cocaine in sample to be checked.
2. according to claim 1 based on rolling circle amplification and the fit method to cocaine Electrochemical Detection of supermolecule, it is characterized in that: in described step 1, gold electrode polishing is the alumina powder of 0.05 μ m by granularity, and described Piranha solution is dense H
2sO
4: H
2o
2volume ratio is the solution of 3:1.
3. according to claim 1 based on rolling circle amplification and the fit method to cocaine Electrochemical Detection of supermolecule, it is characterized in that: the fit fragment Co3S sequence of described marking sulfhydryl is: 5 '-GGGAGTCAAGAACGAAAAAAAA-thiol-(CH
2)
3biotin labeled fit fragment Co3B sequence is: 5 '-TTCGTTCTTCAATGAAGTGGGACGACAAAAAAA-biotin, detector probe sequence is: 5 '-Biotin-AAAAAAGCGCAGAATGGT, while making cyclized DNA, primed DNA sequence used is: 5 '-Biotin-AAAAAAAAAAAACAGGGCTGGGCATAGAAGTCAGGGCAGAGA, treats that cyclisation template DNA sequence is: 5 '-P-TATGCCCAGCCCTGTAAGATGAAGATAGCGCAGAATGGTCGGATTCTCAACTCG TATCTGCCCTGACTTC.
4. according to claim 1 based on rolling circle amplification and the fit method to cocaine Electrochemical Detection of supermolecule, it is characterized in that: described step 3 to the flushing of gold electrode in 7 completes in accordance with the following steps: first with rinsing three times containing 0.005% polysorbas20 Tris-HCl damping fluid, and then with Tris-HCl damping fluid flushing three times; Tris-HCl damping fluid is for containing 20mM Tris, 0.1M sodium chloride, and 5mM magnesium chloride, pH 7.4.
5. according to claim 1 based on rolling circle amplification and the fit method to cocaine Electrochemical Detection of supermolecule, it is characterized in that: the salmon sperm dna that the salmon sperm dna in described step 3 and 2%BSA mixed solution are 10mg/ml by concentration and 2%BSA volume ratio are 1:80 mixed preparing.
6. according to claim 1 based on rolling circle amplification and the fit method to cocaine Electrochemical Detection of supermolecule, it is characterized in that: the preparation method of the testing sample solution in described step 4 is: urine is centrifugal, get supernatant, then with the 1:3 dilution by volume of PB damping fluid; Described step 4), 5), 6) in PB damping fluid for containing 5mM potassium chloride, 5mM magnesium chloride, 40 μ M TCEP, 0.1M phosphate, pH 7.4.
7. according to claim 1 based on rolling circle amplification and the fit method to cocaine Electrochemical Detection of supermolecule, it is characterized in that: in the rolling circle amplification reactant liquor in described step 7, contain 33mM Tris, 10mM magnesium chloride, 66mM potassium chloride, 1mM dithiothreitol (DTT), 0.1% Tween-20, pH 7.9.
8. according to claim 1ly based on rolling circle amplification and the fit method to cocaine Electrochemical Detection of supermolecule, it is characterized in that: the hybridization solution in described step 8 is 2 * SSC damping fluid, contain 0.3M sodium chloride, 0.03M trisodium citrate, pH 7.4; DEA damping fluid is for containing 0.1M diethanolamine, 1mM magnesium chloride, and 100mM potassium chloride, pH 9.6.
9. according to claim 1 based on rolling circle amplification and the fit method to cocaine Electrochemical Detection of supermolecule, it is characterized in that: the concentration of the streptavidin solution of preparing with PB damping fluid in described step 5 is identical with the cyclized DNA concentration in step 6, and the amount adding is also identical.
10. according to claim 1 based on rolling circle amplification and the fit method to cocaine Electrochemical Detection of supermolecule, it is characterized in that: the range of linearity that cocaine detects is 10～500nM, and sensitivity is 9.26nM.
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| CN105510411B (en) * | 2015-09-14 | 2018-05-08 | 陕西师范大学 | The detection method of single cancer cell is realized in interaction based on cell and microelectrode |
| CN106198952A (en) * | 2016-07-01 | 2016-12-07 | 清华大学 | A kind of suppress the nucleic acid molecules enclosure method to sensing interface non-specific adsorption |
| CN107287298A (en) * | 2017-06-26 | 2017-10-24 | 同济大学 | A kind of electrochemical sensing method that DNA damage is detected based on micro-gap array electrode/rolling circle amplification |
| CN107287298B (en) * | 2017-06-26 | 2020-12-11 | 同济大学 | Electrochemical sensing method for detecting DNA damage based on micro-gap array electrode/rolling circle amplification technology |
| CN108148810A (en) * | 2017-12-14 | 2018-06-12 | 临沂大学 | A kind of aptamer and the RNA films of luminol-gold nanoparticle functionalization and its preparation method and application |
| CN108148810B (en) * | 2017-12-14 | 2020-06-12 | 临沂大学 | Aptamer and luminol-gold nanoparticle functionalized RNA membrane and preparation method and application thereof |
| CN111175365A (en) * | 2020-01-20 | 2020-05-19 | 重庆医科大学 | Electrochemical sensor for detecting exosome miRNA (micro ribonucleic acid), and preparation and application thereof |
| CN111175365B (en) * | 2020-01-20 | 2022-03-25 | 重庆医科大学 | Electrochemical sensor for detecting exosome miRNA (micro ribonucleic acid), and preparation and application thereof |
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