CN109060903A - The preparation method of lead ion detection method probe integrated functionality nucleic acid modified electrode in lake water water sample - Google Patents

The preparation method of lead ion detection method probe integrated functionality nucleic acid modified electrode in lake water water sample Download PDF

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Publication number
CN109060903A
CN109060903A CN201811265189.1A CN201811265189A CN109060903A CN 109060903 A CN109060903 A CN 109060903A CN 201811265189 A CN201811265189 A CN 201811265189A CN 109060903 A CN109060903 A CN 109060903A
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electrode
buffer solution
nucleic acid
probe
modified electrode
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申晓萍
王超颖
洪华
张爱平
施筱迪
王海天
洪昱
王红卫
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Nantong Entry-Exit Inspection And Quarantine Bureau Of People's Republic Of
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Nantong Entry-Exit Inspection And Quarantine Bureau Of People's Republic Of
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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
    • G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
    • G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
    • G01N27/28—Electrolytic cell components
    • G01N27/30—Electrodes, e.g. test electrodes; Half-cells
    • G01N27/327—Biochemical electrodes, e.g. electrical or mechanical details for in vitro measurements
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
    • G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
    • G01N27/28—Electrolytic cell components
    • G01N27/30—Electrodes, e.g. test electrodes; Half-cells
    • G01N27/327—Biochemical electrodes, e.g. electrical or mechanical details for in vitro measurements
    • G01N27/3275—Sensing specific biomolecules, e.g. nucleic acid strands, based on an electrode surface reaction
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
    • G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
    • G01N27/28—Electrolytic cell components
    • G01N27/30—Electrodes, e.g. test electrodes; Half-cells
    • G01N27/333—Ion-selective electrodes or membranes
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
    • G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
    • G01N27/416—Systems
    • G01N27/48—Systems using polarography, i.e. measuring changes in current under a slowly-varying voltage

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  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Molecular Biology (AREA)
  • Physics & Mathematics (AREA)
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  • Electrochemistry (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
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  • Spectroscopy & Molecular Physics (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

The invention discloses a kind of preparation methods of lead ion detection method probe integrated functionality nucleic acid modified electrode in lake water water sample, and gold nanoparticle modified electrode is placed in self assembly about 12 hours in the buffer solution added with 1 μM of mercapto-modified 17E DNAzyme；The buffer solution is the 20mM Tris-HCl solution of pH7.5, NaCl containing 140mM and 5mM KCl；It takes out electrode to be cleaned with the buffer solution, then electrode is placed in the above-mentioned buffer solution added with 1mM sulfydryls hexanol and is passivated, cleaned with same buffer solution and remove unassembled mercaptoethanol；Finally electrode is transferred in the substrate 17DS solution containing 1 μM and is assembled；The terminal modified ferrocene probe molecule in the 3 ' of the substrate 17DS；Electrode is washed with above-mentioned buffer solution again, obtains the integrated functional nucleic acid modified electrode of probe.The present invention is selectively good, sensitive, easy, quick.

Description

Lead ion detection method probe integrated functionality nucleic acid modified electrode in lake water water sample Preparation method
The application is application number: 201610372050.1, the applying date: 2016-05-31, title " the integrated function core of probe The divisional application of sour modified electrode Direct Electrochemistry detection lead ion method ".
Technical field
The present invention relates to the functional nucleic acid modified electrode Direct Electrochemistries that a kind of probe integrates to detect lead ion method.
Background technique
Lead ion is a kind of common harmful heavy metal ions, and monitoring/detection has human health and environmental monitoring Highly important meaning.In the past few decades, people are had been developed such as Atomic absorption/emission spectrum, fluorescence spectrum, matter A variety of methods such as spectrum, photonic crystal are monitored/detect to lead ion.In recent years, Lu Yi research group in the U.S. by sieving in vitro Selecting technology obtains the functional nucleic acid 17E DNAzyme sensitive to lead ion, they are identified using 17E DNAzyme as lead ion Element uses gold nanoparticle as sensing element, develops colorimetric sensor detection lead ion (Liu, J.; Lu, Y., Nature Protocols 2006, 1, 246-252.).Wang group is based on 17E DNAzyme and and double-stranded DNA fluorescence Molecule chelating dyes Picogreen has developed label-free fluorescent optical sensor detection lead ion (Zhang, L.; Han, B.; Li, T.; Wang, E., Chemical Communications 2011, 47, 3099–3101.).Then, Yin group Electrochemical luminescence sensor detection lead ion (Gao, A. are developed based on 17E DNAzyme; Tang, C.; He, X.; Yin, X., Analyst, 2013, 138, 263–268).Therefore the functional nucleic acid 17E DNAzyme of lead ion sensitivity is utilized Detection lead ion receives the extensive concern of domestic and international expert.
Since the detection of fluorescence detection, ultraviolet detection and electrochemical luminescence requires huger, valuable detecting instrument, Therefore these methods are not suitable for the occasion of some simpler and cruder yards field work in one's power.Electrochemical detection method due to Have many advantages, such as high sensitivity, it is easy to operate it is quick, at low cost, low energy consumption, is easy to instrumentation, thus day by researcher Benefit favor.
Summary of the invention
The purpose of the present invention is to provide a kind of selectivity, and the integrated functional nucleic acid of good, sensitive, easy, quick probe is repaired It adorns electrode Direct Electrochemistry and detects lead ion method.
The technical solution of the invention is as follows:
A kind of functional nucleic acid modified electrode Direct Electrochemistry detection lead ion method that probe is integrated, it is characterized in that: by probe collection At functional nucleic acid modified electrode immerse added with buffer 20 ~ 30 minutes of lead ion to be measured, take out washing and 0.1 M's NaClO4DPV measurement is carried out in electrolyte solution, according to the DPV peak current signal for scanning record within the scope of 0.1~0.6V Change rate calculates the plumbum ion concentration in sample；The buffer is pH 7.5,20 containing 140 mM NaCl and 5 mM KCl MM Tris-HCl solution；
The integrated functional nucleic acid modified electrode of the probe is prepared by following methods: gold nanoparticle modified electrode is placed in Self assembly 10 ~ 18 hours in buffer solution added with 1 μM of mercapto-modified 17E DNAzyme；The buffer be pH 7.5, The 20 mM Tris-HCl solution containing 140 mM NaCl and 5 mM KCl；It takes out electrode to be cleaned with the buffer solution, then will be electric Pole is placed in the above-mentioned buffer solution added with 1mM sulfydryls hexanol and is passivated 1-3 hours, cleans removing not with same buffer solution The mercaptoethanol of assembling；Finally electrode is transferred in the substrate 17DS solution containing 1 μM and is assembled 10 ~ 18 hours；The substrate The terminal modified ferrocene probe molecule in the 3 ' of 17DS；Electrode is washed 2-3 times with above-mentioned buffer solution again, obtains the integrated function of probe Nucleic acid modified electrode.
The integrated functional nucleic acid modified electrode of probe immerses buffer 25 minutes added with lead ion to be measured.
Gold nanoparticle modified electrode is placed in the buffer solution added with 1 μM of mercapto-modified 17E DNAzyme from group It fills 12 hours.
Finally electrode is transferred in the substrate 17DS solution containing 1 μM and is assembled 12 hours.
The invention has the benefit that
1. the identification molecule that the present invention is detected using functional nucleic acid 17E DNAzyme as lead ion, selectivity is good, to environment It is without secondary pollution.
2. detected using electrochemical method, have high sensitivity, instrument it is simple, it is easy to operate quickly, low cost and low energy The advantages that consumption.
3. this method is integrated into electrode surface, is not necessarily in detection process using the method for probe and substrate chain 17DS integration Additional electrochemical probe reagent is added in electrolyte solution, the degree of integration of electrode is high, functional, and detection convenience mentions It is high.
Detailed description of the invention
Present invention will be further explained below with reference to the attached drawings and examples.
Fig. 1 is the DPV curvilinear motion figure in the present invention to the modified electrode measured during lead at different concentrations ion detection.
Fig. 2 is the standard working curve figure that modified electrode of the invention detects lead ion.
Specific embodiment
(1) preparation of the integrated functional nucleic acid modified electrode of probe
Gold nanoparticle modified electrode is placed in self assembly 12 in the buffer solution added with 1 μM of mercapto-modified 17E DNAzyme Hour；The buffer is pH 7.5, the 20 mM Tris-HCl solution containing 140 mM NaCl and 5 mM KCl；Take out electrode It is cleaned with the buffer solution, then electrode is placed in the above-mentioned buffer solution added with 1mM sulfydryls hexanol and is passivated 2 hours, used Same buffer solution cleaning removes unassembled mercaptoethanol；Electrode is finally transferred to the substrate 17DS solution containing 1 μM Middle assembling 12 hours；The terminal modified ferrocene probe molecule in the 3 ' of the substrate 17DS；Electrode 3 is washed with above-mentioned buffer solution again It is secondary, obtain the integrated functional nucleic acid modified electrode of probe.
(2) in lake water water sample lead ion detection
The modified electrode prepared is immersed in a series of buffer solutions added with the lead ion standard water sample of various concentration and is soaked After steeping 25 min, with above-mentioned buffer solution cleaning electrode 2 times；With the NaClO of 0.1 M4As supporting electrolyte solution, carry out DPV sweep test, test parameter are as follows: adjustment time is 50 ms, and interval time is 0.5 s, and modulated amplitude is 50 mV, step electricity Gesture is 5 mV, and scanning range is 0.1~0.6V.Using DPV peak current as ordinate, plumbum ion concentration is abscissa, draws standard Working curve, as shown in Figure 2.
Using lake water as actual sample, lake water after active carbon decoloring with 0.22 μm of membrane filtration, then with 15000 Turn/min centrifugation after, take supernatant to be measured.The 50 μ L lake water water sample handled well is added in above-mentioned buffer solution, is pressed According to said determination step operation, its DPV peak current is measured, according to DPV peak current change rate (I0-I)/I0, intended by standard curve It is 327 pM that the linear equation of conjunction, which can calculate plumbum ion concentration in water sample,.

Claims (1)

1. preparation method of the lead ion detection method with probe integrated functionality nucleic acid modified electrode, feature in a kind of lake water water sample Be: lead ion detection method is in the lake water water sample: the modified electrode prepared being immersed in a series of added with various concentration Lead ion standard water sample buffer solution in impregnate 25 min after, with above-mentioned buffer solution cleaning electrode；With 0.1 M's NaClO4As supporting electrolyte solution, DPV sweep test, test parameter are carried out are as follows: adjustment time is 50 ms, interval time For 0.5 s, modulated amplitude is 50 mV, and step potential is 5 mV, and scanning range is 0.1~0.6V；It is vertical sit with DPV peak current Mark, plumbum ion concentration is abscissa, draws standard working curve；
Using lake water as sample, lake water after active carbon decoloring with 0.22 μm of membrane filtration, then with 15000 turns/min from After the heart, supernatant is taken to be measured；The 50 μ L lake water water sample handled well is added in above-mentioned buffer solution, according to above-mentioned survey Determine step operation, measure its DPV peak current, according to DPV peak current change rate (I0-I)/I0, by the linear of standard curve fit Equation can calculate plumbum ion concentration in water sample；
The preparation method of the integrated functional nucleic acid modified electrode of the probe is: gold nanoparticle modified electrode is placed in added with 1 Self assembly about 12 hours in the buffer solution of μM mercapto-modified 17E DNAzyme；The buffer solution is pH 7.5, containing 140 The 20 mM Tris-HCl solution of mM NaCl and 5 mM KCl；It takes out electrode to be cleaned with the buffer solution, then electrode is placed in It is passivated into the above-mentioned buffer solution added with 1mM sulfydryls hexanol, is cleaned with same buffer solution and remove unassembled sulfydryl second Alcohol；Finally electrode is transferred in the substrate 17DS solution containing 1 μM and is assembled；3 ' the terminal modified ferrocene of the substrate 17DS Probe molecule；Electrode is washed with above-mentioned buffer solution again, obtains the integrated functional nucleic acid modified electrode of probe.
CN201811265189.1A 2016-05-31 2016-05-31 The preparation method of lead ion detection method probe integrated functionality nucleic acid modified electrode in lake water water sample Pending CN109060903A (en)

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CN102375020A (en) * 2010-08-19 2012-03-14 华东师范大学 Method for detecting mercury ions through gold electrode modified by polythymidine DNA
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Application publication date: 20181221