CN115060777B - Rate electrochemical aptamer sensor for simultaneously detecting malathion and omethoate, and preparation method and application thereof - Google Patents
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Abstract
The invention relates to the technical field of electrochemical sensing, in particular to a ratio electrochemical aptamer sensor for simultaneously detecting malathion and omethoate, a preparation method and application thereof, which are obtained by sequentially assembling a CNHs/AQ/AuNPs composite material, hDNA, MCH, MB-Apt1 and Fc-Apt2 on a glassy carbon electrode. When MAL or OMT is present, its specific binding to the aptamer causes MB-Apt1 or Fc-Apt2 to be peeled off the electrode surface, resulting in a ratio signal I MB /I AQ Or I Fc /I AQ The ratio of MAL and OMT is reduced, and simultaneous detection is realized. The detection linear range of the MAL is 3 pg/mL-3 ng/mL, and the detection limit is 1.3pg/mL; the linear detection range of OMT is 10 pg/mL-10 ng/mL, and the detection limit is 2.8pg/mL. The sensor has the advantages of high sensitivity, good selectivity, high accuracy and the like.
Description
Technical Field
The invention relates to the technical field of electrochemical sensing, in particular to a ratio electrochemical aptamer sensor for simultaneously detecting malathion and omethoate.
Background
The organophosphorus pesticides are a class of organophosphorus pesticides commonly used in agricultural production, and are mainly used for preventing, controlling or eliminating diseases, insect pests and weeds so as to improve the quality and yield of crops. Malathion (MAL) and Omethoate (OMT) are two commonly used organophosphorus pesticides, which are widely used by farmers in various countries of the world due to their advantages of broad spectrum, high efficiency, low price, etc.
The organophosphorus pesticide has high neurotoxicity, can inhibit activity of acetylcholinesterase in nervous system even at low concentration, can accumulate acetylcholinesterase, and can cause a series of neurotoxic symptoms, and can die due to pulmonary edema, cerebral edema and respiratory paralysis when serious. Due to the widespread use of organophosphorus pesticides, their residue in agricultural products has caused serious human health and environmental problems. Therefore, development of effective detection techniques is required to achieve efficient detection analysis of MAL and OMT in agricultural products.
At present, simultaneous detection technologies for MAL and OMT mainly comprise an enzyme-linked immunosorbent assay, gas chromatography, high performance liquid chromatography-mass spectrometry and the like, but the detection technologies have the defects of expensive instrument, long time consumption, complex operation and the like, and a simple, convenient, rapid and efficient detection means needs to be developed.
The electrochemical sensing technology has the advantages of low cost, quick response, simple operation and the like, and is expected to be a very promising method for simultaneously detecting various pesticides with high efficiency. Currently, electrochemical aptamer sensors for single pesticides already have good sensitivity and selectivity, but the detection accuracy thereof needs to be further improved.
Therefore, the development of a technical scheme capable of realizing high-sensitivity and high-precision simultaneous detection of MAL and OMT has important research significance.
Disclosure of Invention
The invention aims to provide a ratio electrochemical aptamer sensor for simultaneously detecting malathion and omethoate, a preparation method and application thereof.
The ratio electrochemical aptamer sensor based on the ratio strategy has double response signals aiming at the target object, and the concentration of the target object is quantified by measuring the ratio of the double signals, so that the accuracy of the electrochemical sensor can be effectively improved. According to the invention, a carbon nanohorn/anthraquinone-2-carboxylic acid/gold nanoparticle composite material (CNHs/AQ/AuNPs), hairpin DNA (hDNA), mercapto hexanol (MCH), methylene blue marked malathion aptamer (MB-Apt 1) and ferrocene marked omethoate aptamer (Fc-Apt 2) are sequentially assembled on a glassy carbon electrode to construct a ratio electrochemical aptamer sensor, so that the high-efficiency simultaneous detection of malathion and omethoate is realized.
Specifically, the invention provides the following technical scheme:
a method for preparing a ratio electrochemical aptamer sensor for simultaneously detecting malathion and omethoate, comprising the following steps:
(1) Sequentially polishing the glassy carbon electrode GCE with aluminum oxide powder with different particle sizes, sequentially ultrasonically treating the glassy carbon electrode GCE in ethanol and pure water to remove surface residues, and airing the glassy carbon electrode GCE at room temperature;
(2) Dripping the prepared carbon nanohorn/anthraquinone-2-carboxylic acid/gold nanoparticle composite material, namely CNHs/AQ/AuNPs, onto the electrode treated in the step (1), and airing at room temperature;
(3) Modifying hairpin DNA, i.e. hDNA, onto the electrode treated in the step (2), storing for a period of time at room temperature, and fixing the hDNA on the surface of the electrode by utilizing gold-sulfur bonds;
(4) Modifying sulfhydryl hexanol (MCH) on the electrode treated in the step (3), and incubating for a period of time under the room temperature condition to block non-specific binding sites of gold;
(5) The methylene blue marked malathion aptamer is marked as MB-Apt1, modified on the electrode treated in the step (4), reacted for a period of time under a certain temperature condition, and the MB-Apt1 is assembled on the electrode by utilizing the complementary hybridization of the methylene blue marked malathion aptamer and hDNA;
(6) And (3) modifying the ferrocene-labeled omethoate aptamer which is marked as Fc-Apt2 on the electrode treated in the step (5), reacting for a period of time under a certain temperature condition, and further assembling the Fc-Apt2 on the electrode by utilizing complementary hybridization to obtain the ratio electrochemical aptamer sensor which is marked as MB-Apt1 and Fc-Apt2/MCH/hDNA/CNHs/AQ/AuNPs/GCE.
Preferably, in the step (1), the diameter of the glassy carbon electrode is 3mm; the particle size of the aluminum oxide powder used was 0.3 μm and 0.05 μm in this order.
Preferably, in the step (2), the preparation steps of the CNHs/AQ/AuNPs composite material are as follows: firstly dispersing 1-10 mg of carbon nanohorn in 2-20 mL of water, and stirring for 0.5-2 hours at room temperature to obtain solution A; adding 0.1-1 mg of anthraquinone-2-carboxylic Acid (AQ) into the solution A, stirring for 4-12 hours at room temperature, centrifuging and washing for 3 times, and then re-dispersing the precipitate into 2-20 mL of pure water to obtain a solution B; adding 0.05-0.5 mL of HAuCl with the concentration of 30-60 mM into the solution B 4 Stirring the solution for 0.5 to 1 hour at room temperature, adding 0.5 to 5mL of ascorbic acid solution with the concentration of 2 to 20mM, continuously stirring for 4 to 12 hours at room temperature, centrifuging, washing and drying to obtain the CNHs/AQ/AuNPs composite material; dispersing the obtained CNHs/AQ/AuNPs composite material in 2-20 mL of pure water to obtain CNHs/AQ/AuNPs solution.
Preferably, in the step (2), the CNHs/AQ/AuNPs solution is used in an amount of 4-8 mu L and the concentration is 0.05-5 mg/mL.
Preferably, in the step (3), the amount of the hDNA is 4-8 mu L and the concentration is 1-8 mu M; the storage time is 4-12 hours under the room temperature condition.
Preferably, in the step (4), the MCH is used in an amount of 4 to 8. Mu.L and the concentration is 0.01 to 10mM; the incubation time is 0.5-2 hours under the room temperature condition.
Preferably, in the step (5), the MB-Apt1 is used in an amount of 4-8 mu L and the concentration is 1-8 mu M; the temperature is 37 ℃ and the reaction time is 1-2 hours.
Preferably, in the step (6), the amount of the Fc-Apt2 is 4-8 mu L and the concentration is 1-8 mu M; the temperature is 37 ℃ and the reaction time is 1-2 hours.
The working method for detecting malathion and omethoate by using the sensor prepared by the invention specifically comprises the following steps:
(1) The prepared multiple sensors MB-Apt1&Fc-Apt2/MCH/hDNA/CNHs/AQ/AuNPs/GCE are sequentially added with malathion and omethoate standard solutions with different concentrations in a dropwise manner, and each sensor corresponds to one concentration; after incubation for a certain period of time at room temperature, the incubation was performed using Tris-HCl (ph=7.4) solutionFlushing; in a three-electrode system, the washed electrode is used as a working electrode, an Ag/AgCl (saturated KCl) electrode is used as a reference electrode, a platinum wire is used as a counter electrode, a phosphate buffer solution (pH=7.4) is used as an electrolyte, an electrochemical Alternating Current Voltammogram (ACV) is scanned, and a ratio signal I is measured MB /I AQ And I Fc /I AQ Establishing ratio signals I corresponding to malathion and omethoate concentrations respectively MB /I AQ And I Fc /I AQ Is a standard linear curve of (2);
wherein I is MB Is MB oxidation current, I Fc For Fc oxidation current, I AQ Is AQ oxidation current;
(2) Drop-adding sample solution to be measured into sensor MB-Apt1&Fc-Apt2/MCH/hDNA/CNHs/AQ/AuNPs/GCE, after incubation for a certain period of time at room temperature, rinsing with Tris-HCl (pH=7.4) solution; in a three-electrode system, the washed electrode is used as a working electrode, an Ag/AgCl (saturated KCl) electrode is used as a reference electrode, a platinum wire is used as a counter electrode, a phosphate buffer solution (pH=7.4) is used as an electrolyte, an electrochemical Alternating Current Voltammogram (ACV) is scanned, and a ratio signal I is measured MB /I AQ And I Fc /I AQ Substituting the standard linear curve in the step (1) to realize high-precision simultaneous detection of malathion and omethoate of the sample to be detected.
Preferably, in step (1), the concentration of malathion ranges from 3×10 -12 ～3×10 -9 g/mL, the concentration range of omethoate is 1 multiplied by 10 -11 ～1×10 -8 g/mL。
Preferably, in the steps (1) and (2), the incubation is performed for a period of time ranging from 0.5 to 2 hours at room temperature.
The working principle of the sensor prepared by the invention is as follows:
first, sequentially modified CNHs/AQ/AuNPs, hDNA, MCH, MB-Apt1 and Fc-Apt2 are assembled on the surface of the glassy carbon electrode to form a double-stranded DNA structure based on hairpin DNA, and a large MB oxidation current is generated (I MB ) Fc oxidation current (I) Fc ) And AQ oxidation current (I) AQ ). When malathion or omethoate is present, its specific binding to the aptamer results in the production of MB-Apt1 or Fc-Apt2 fromElectrode surface delamination, leading to I MB And I Fc Decrease, and I AQ Remain unchanged. Thus, the ratio signal I can be measured MB /I AQ And I Fc /I AQ Realizes the simultaneous detection of target substances malathion and omethoate.
Compared with the prior art, the invention has the beneficial effects that:
(1) The CNHs/AQ/AuNPs composite material prepared in the invention can generate an internal standard signal I AQ And amplifies the target response signal I MB And I Fc 。
(2) The invention constructs a sensing interface by utilizing the specific complementary hybridization of the hairpin DNA and 2 kinds of aptamer, and the 2 kinds of targets can not generate cross influence.
(3) The electrochemical aptamer sensor for simultaneously detecting the malathion and the omethoate, which is prepared by the invention, has the characteristics of high sensitivity, good selectivity, high accuracy and the like, and has potential application prospect in the field of agricultural product quality detection.
Drawings
Fig. 1 is a schematic diagram of the construction and detection process of the ratio electrochemical sensor.
FIG. 2 (A) shows ACV responses corresponding to MAL and OMT at different concentrations, wherein MAL concentrations are 3×10 in order -12 ,1×10 -11 ,3×10 -11 ,1×10 -10 ,3×10 -10 ,1×10 -9 And 3X 10 -9 g/mL; OMT concentration of 1X 10 in order -11 ,3×10 -11 ,1×10 -10 ,3×10 -10 ,1×10 -9 ,3×10 -9 And 1X 10 -8 g/mL; (B) For ratio signal I MB /I AQ And I Fc /I AQ Standard linear curves between the MAL and OMT concentration logarithms, respectively.
Detailed Description
The following description of the embodiments of the present invention will be made clearly and completely with reference to the accompanying drawings, in which it is apparent that the embodiments described are only some embodiments of the present invention, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
A preparation method of a ratio electrochemical aptamer sensor for simultaneously detecting malathion and omethoate is provided, wherein the construction and detection processes of the ratio electrochemical aptamer sensor are shown in the accompanying figure 1.
The method specifically comprises the following steps:
(1) Sequentially polishing the glassy carbon electrode GCE with the diameter of 3mm by using aluminum oxide powder with the diameter of 0.3 mu m and aluminum oxide powder with the diameter of 0.05 mu m, sequentially ultrasonically treating the mixture in ethanol and water to remove surface residues, and airing the mixture at room temperature;
(2) Dropping 6 mu L of CNHs/AQ/AuNPs solution with the concentration of 1mg/mL onto the electrode treated in the step (1), and airing at room temperature;
the preparation method of the CNHs/AQ/AuNPs solution comprises the following steps:
firstly, dispersing 5mg of carbon nanohorn in 10mL of water, and stirring for 0.5 hour at room temperature to obtain solution A; adding 0.35mg of anthraquinone-2-carboxylic Acid (AQ) into the solution A, stirring for 12 hours at room temperature, centrifuging and washing for 3 times, and then re-dispersing the precipitate into 10mL of pure water to obtain a solution B; to solution B was added 0.2mL of HAuCl at 60mM 4 Stirring the solution for 0.5 hour at room temperature, adding 2mL of ascorbic acid solution with the concentration of 10mM, continuously stirring for 8 hours at room temperature, centrifuging, washing and drying to obtain a CNHs/AQ/AuNPs composite material; and dispersing the obtained CNHs/AQ/AuNPs composite material in 5mL of pure water to obtain CNHs/AQ/AuNPs solution.
(3) 6 mu L of hDNA with the concentration of 2 mu M is modified on the electrode treated in the step (2), and the electrode is placed for 8 hours at room temperature, and the hDNA is fixed on the surface of the electrode by gold-sulfur bonds;
(4) 6. Mu.L of 1mM Mercaptohexanol (MCH) was modified on the electrode treated in step (3) and incubated at room temperature for 1 hour to block the gold non-specific binding sites;
(5) Modifying 6 mu L of methylene blue marked malathion aptamer (MB-Apt 1) with the concentration of 2 mu M on the electrode treated in the step (4), reacting for 1 hour at 37 ℃, and assembling MB-Apt1 on the electrode by utilizing complementary hybridization of the methylene blue marked malathion aptamer and hDNA;
(6) 6 mu L of ferrocene-labeled omethoate aptamer (Fc-Apt 2) with concentration of 2 mu M is modified on the electrode treated in the step (5), and reacted at 37 ℃ for 1 hour, and the Fc-Apt2 is further assembled on the electrode by utilizing complementary hybridization to obtain the ratio electrochemical aptamer sensor which is marked as MB-Apt1& Fc-Apt2/MCH/hDNA/CNHs/AQ/AuNPs/GCE.
The prepared sensor MB-Apt1&Fc-Apt2/MCH/hDNA/CNHs/AQ/AuNPs/GCE are sequentially added with malathion and omethoate standard solutions with different concentrations in a dropwise manner, wherein the concentration of MAL is sequentially 3 multiplied by 10 -12 ,1×10 -11 ,3×10 -11 ,1×10 -10 ,3×10 -10 ,1×10 -9 And 3X 10 -9 g/mL, OMT concentration of 1X 10 in order -11 ,3×10 -11 ,1×10 -10 ,3×10 -10 ,1×10 -9 ,3×10 -9 And 1X 10 -8 g/mL; after incubation for 40 min at room temperature, rinsing was performed using Tris-HCl (ph=7.4) solution. In a three-electrode system, the washed electrode is used as a working electrode, an Ag/AgCl (saturated KCl) electrode is used as a reference electrode, a platinum wire is used as a counter electrode, a phosphate buffer solution (pH=7.4) is used as an electrolyte, an electrochemical Alternating Current Voltammogram (ACV) is scanned, and a ratio signal I is measured MB /I AQ And I Fc /I AQ Establishing ratio signals I corresponding to malathion and omethoate concentrations respectively MB /I AQ And I Fc /I AQ Is a standard linear curve of (2).
From FIG. 2 (A), it can be seen that as the MAL and OMT concentrations increase, I MB And I Fc The values of (2) respectively gradually decrease.
From FIG. 2 (B), it can be seen that the ratio I MB /I AQ A standard linear curve between MAL concentration logarithm is I MB /I AQ ＝-0.161-0.122LogC MAL [g/mL](R 2 =0.997), the linear range is 3pg/mL to 3ng/mL, and the detection limit is 1.3pg/mL; ratio I Fc /I AQ Standard linear curve between OMT concentration logarithm is I Fc /I AQ ＝-0.279-0.098LogC OMT [g/mL](R 2 =0.998), the linear range is 10pg/mL to 10ng/mL, and the detection limit is 2.8pg/mL.
The obtained sensor is applied to the marking detection of malathion and omethoate with different concentrations in lettuce as actual samples, and the detection process is as follows:
drop-adding sample solution to sensor MB-Apt1&Fc-Apt2/MCH/hDNA/CNHs/AQ/AuNPs/GCE, after incubation for 40 min at room temperature, was rinsed with Tris-HCl (pH=7.4) solution. In a three-electrode system, the washed electrode is used as a working electrode, an Ag/AgCl (saturated KCl) electrode is used as a reference electrode, a platinum wire is used as a counter electrode, a phosphate buffer solution (pH=7.4) is used as an electrolyte, an electrochemical Alternating Current Voltammogram (ACV) is scanned, and a ratio signal I is measured FB /I AQ And I Fc /I AQ Substituting the standard linear curve I MB /I AQ ＝-0.161-0.122LogC MAL [g/mL]I Fc /I AQ ＝-0.279-0.098LogC OMT [g/mL]And calculating to obtain concentration information of malathion and omethoate in lettuce.
To verify the accuracy of the constructed sensors, the same lettuce samples were tested using high performance liquid chromatography-mass spectrometry (HPLC-MS). The results of the two methods are shown in Table 1.
TABLE 1 detection results of MAL and OMT in lettuce samples respectively by the sensor constructed in the present invention and high performance liquid chromatography-mass spectrometry
The results in table 1 show that the sensor constructed by the invention can realize high-sensitivity and high-precision simultaneous detection of malathion and omethoate.
Although embodiments of the present invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made therein without departing from the principles and spirit of the invention, the scope of which is defined in the appended claims and their equivalents.
Claims (10)
1. A method for preparing a ratio electrochemical aptamer sensor for simultaneously detecting malathion and omethoate, which is characterized by comprising the following steps:
(1) Sequentially polishing the glassy carbon electrode GCE with aluminum oxide powder with different particle sizes, sequentially ultrasonically treating the glassy carbon electrode GCE in ethanol and pure water to remove surface residues, and airing the glassy carbon electrode GCE at room temperature;
(2) Dripping the prepared carbon nanohorn/anthraquinone-2-carboxylic acid/gold nanoparticle composite material, namely CNHs/AQ/AuNPs, onto the electrode treated in the step (1), and airing at room temperature;
(3) Modifying hairpin DNA, i.e. hDNA, onto the electrode treated in the step (2), storing for a period of time at room temperature, and fixing the hDNA on the surface of the electrode by utilizing gold-sulfur bonds;
(4) Modifying the sulfhydryl hexanol MCH on the electrode treated in the step (3), and incubating for a period of time under the room temperature condition so as to block the non-specific binding site of gold;
(5) Modifying methylene blue marked malathion aptamer MB-Apt1 on the electrode treated in the step (4), reacting for a period of time under a certain temperature condition, and assembling MB-Apt1 on the electrode by utilizing complementary hybridization of the methylene blue marked malathion aptamer MB-Apt1 and hDNA;
(6) And (3) modifying the ferrocene-labeled omethoate aptamer Fc-Apt2 on the electrode treated in the step (5), reacting for a period of time under a certain temperature condition, and further assembling the Fc-Apt2 on the electrode by utilizing complementary hybridization to obtain the ratio electrochemical aptamer sensor, which is marked as MB-Apt1& Fc-Apt2/MCH/hDNA/CNHs/AQ/AuNPs/GCE.
2. The method for preparing the electrochemical aptamer sensor for simultaneously detecting the ratio of malathion and omethoate according to claim 1, wherein the method comprises the following steps: in the step (1), the diameter of the glassy carbon electrode is 3mm; the particle size of the aluminum oxide powder used was 0.3 μm and 0.05 μm in this order.
3. The method for preparing the electrochemical aptamer sensor for simultaneously detecting the ratio of malathion and omethoate according to claim 1, wherein the method comprises the following steps: in the step (2), the preparation method of the carbon nanohorn/anthraquinone-2-carboxylic acid/gold nanoparticle composite material comprises the following steps:
(1) Dispersing 1-10 mg of carbon nanohorn in 2-20 mL of water, and stirring for 0.5-2 hours at room temperature to obtain solution A;
(2) Adding 0.1-1 mg of anthraquinone-2-carboxylic acid into the solution A, stirring for 4-12 hours at room temperature, centrifuging and washing for 3 times, and then re-dispersing the precipitate into 2-20 mL of pure water to obtain a solution B;
(3) Adding 0.05-0.5 mL of HAuCl with the concentration of 30-60 mM into the solution B 4 Stirring the solution for 0.5 to 1 hour at room temperature, adding 0.5 to 5mL of ascorbic acid solution with the concentration of 2 to 20mM, continuously stirring for 4 to 12 hours at room temperature, centrifuging, washing and drying to obtain the CNHs/AQ/AuNPs composite material;
(4) Dispersing the CNHs/AQ/AuNPs composite material in 2-20 mL of pure water to obtain CNHs/AQ/AuNPs solution.
4. The method for preparing the electrochemical aptamer sensor for simultaneously detecting the ratio of malathion and omethoate according to claim 1, wherein the method comprises the following steps: in the step (2), the dosage of the CNHs/AQ/AuNPs solution is 4-8 mu L, and the concentration is 0.05-5 mg/mL.
5. The method for preparing the electrochemical aptamer sensor for simultaneously detecting the ratio of malathion and omethoate according to claim 1, wherein the method comprises the following steps: in the step (3), the dosage of the hDNA is 4-8 mu L and the concentration is 1-8 mu M; the storage time is 4-12 hours under the room temperature condition.
6. The method for preparing a ratio electrochemical aptamer sensor for simultaneous detection of malathion and omethoate according to claim 1, wherein in step (4), MCH is used in an amount of 4 to 8 μl and at a concentration of 0.01 to 10mM; the incubation time is 0.5-2 hours under the room temperature condition.
7. The method for preparing the electrochemical aptamer sensor for simultaneously detecting the ratio of malathion and omethoate according to claim 1, wherein the method comprises the following steps: in the step (5), the using amount of MB-Apt1 is 4-8 mu L, and the concentration is 1-8 mu M; the temperature is 37 ℃ and the reaction time is 1-2 hours.
8. The method for preparing the electrochemical aptamer sensor for simultaneously detecting the ratio of malathion and omethoate according to claim 1, wherein the method comprises the following steps: in the step (6), the dosage of the Fc-Apt2 is 4-8 mu L and the concentration is 1-8 mu M; the temperature is 37 ℃ and the reaction time is 1-2 hours.
9. A ratio electrochemical aptamer sensor for simultaneous detection of malathion and omethoate prepared by the method of any one of claims 1-8.
10. The method of operating a ratio electrochemical aptamer sensor for simultaneous detection of malathion and omethoate as claimed in claim 9, comprising the steps of:
(1) The prepared multiple sensors MB-Apt1&Fc-Apt2/MCH/hDNA/CNHs/AQ/AuNPs/GCE are sequentially added with malathion and omethoate standard solutions with different concentrations in a dropwise manner, and each sensor corresponds to one concentration; after incubation for 0.5-2 hours at room temperature, flushing is carried out by using Tris-HCl solution; in a three-electrode system, the washed electrode is used as a working electrode, an Ag/AgCl electrode is used as a reference electrode, a platinum wire is used as a counter electrode, a phosphate buffer solution is used as electrolyte, an electrochemical alternating current volt-ampere curve is scanned, and a ratio signal I is measured MB /I AQ And I Fc /I AQ Establishing ratio signals I corresponding to malathion and omethoate concentrations respectively MB /I AQ And I Fc /I AQ Is a standard linear curve of (2); wherein I is MB Is MB oxidation current, I Fc For Fc oxidation current, I AQ Is AQ oxidation current;
wherein the concentration range of malathion is 3×10 -12 ～3×10 -9 g/mL, the concentration range of omethoate is 1 multiplied by 10 -11 ～1×10 -8 g/mL；
(2) Drop-adding sample solution to be measured into sensor MB-Apt1&After incubating Fc-Apt2/MCH/hDNA/CNHs/AQ/AuNPs/GCE for 0.5-2 hours at room temperature, washing with Tris-HCl solution; in a three-electrode system, the washed electrode is used as a working electrode, an Ag/AgCl electrode is used as a reference electrode, a platinum wire is used as a counter electrode, a phosphate buffer solution is used as electrolyte, an electrochemical alternating current volt-ampere curve is scanned, and a ratio signal I is measured MB /I AQ And I Fc /I AQ Substituting the standard linear curve in the step (1) to realize simultaneous detection of malathion and omethoate of the sample to be detected.
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