Preparation method for ratio electrochemical biosensor for detecting aflatoxin B1
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A kind of aflatoxin and sensor technology, applied in the field of biosensors, to achieve the effect of improving accuracy, wide linear range and high sensitivity
Active Publication Date: 2019-09-06
JIANGSU UNIV
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At present, most studies on ratiometric biosensors use electrochemical signals such as ferrocene (Fc) and methylene blue (MB) to label DNA molecules. Another electrochemical molecule carried by DNA strands to obtain a high-sensitivity ratiometric electrochemical biosensor
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Embodiment 1
[0033] according to figure 1 The preparation process described above, the preparation method of the ratiometric electrochemical biosensor comprises the following steps:
[0034] (1) Preparation of THI-rGO composites:
[0035] First, weigh 1mg THI and dissolve it in ultrapure water to obtain a concentration of 20μg·mL in 50mL -1 THI solution, and serially diluted to 5 μg·mL -1 , stored away from light at room temperature; weigh 3mg rGO and disperse it in 30mL ultrapure water to obtain a concentration of 0.1mg·mL -1 The rGO dispersion solution was treated with a cell pulverizer for 30 min and stored at room temperature until use.
[0036] Then, take 3mL of 5μg·mL -1 THI with 15mL 0.1mg·mL -1 Mix rGO in a 100mL beaker and stir for 12h. After the stirring is over, put the solution in a 10mL centrifuge tube. Under the conditions of rotating speed of 8000rpm and centrifugation time of 15min, centrifuge and wash three times to obtain a precipitate
[0037] Finally, the obtained...
Embodiment 2
[0045] Optimized experimental conditions for the prepared ratiometric electrochemical biosensor for detection of aflatoxin B1:
[0046] The biosensor prepared in Example 1 was placed at 4° C. for 6, 8, 10, 12, and 14 hours to detect Fc electrochemical signals.
[0047] figure 2 (A) AFB1 aptamer incubation time and I Fc Relationship diagram, the incubation time of the aptamer increased from 6h to 10h, I Fc Gradually increased and reached a plateau at 10h. Therefore, 12h was chosen as the optimal incubation time for aptamers.
Embodiment 3
[0049] Optimized experimental conditions for the prepared ratiometric electrochemical biosensor for detection of aflatoxin B1:
[0050] On the basis of the ratio biosensor with the best conditions obtained in Example 2, modify the target AFB1 on the surface of the sensor, such as figure 2 (B) is the target incubation time and I Fc Relationship diagram, the incubation time of the target on the electrode surface is from 0.3h to 1.0h, I Fc The signal gradually decreased, and after 1h I Fc There was no significant change in the signal, so the incubation time of the selected target AFB1 was 1.3h.
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Abstract
The invention belongs to the technical field of biosensors, and particularly relates to a preparation method for a ratio electrochemical biosensor for detecting aflatoxin B1. The composite material ofTHI (thionine) and rGO (reduced graphene oxide) is used as a first beacon molecule, ferrocene carried by AFB1 (Aflatoxin B1) aptamer is used as a second beacon molecule, and chitosan with positive electricity is used for connecting the first beacon molecule with the second beacon molecule through an electrostatic adsorption function and fixing the connected first beacon molecule and second beaconmolecule on the surface of a glassy carbon electrode. In the preparation method, the AFB1 aptamer is introduced, and the high specificity of the ratio electrochemical biosensor toxin AFB1 is improved. Two ends of the aptamer are designed to mark with an Fc signal to realize the amplification of a contrast ratio signal. The detection linear range of the constructed ratio electrochemical biosensoris 0.01ng.mL<-1>-100ng.mL<-1>, and the detection limit is 3.3pg. mL(-1). Through research, the preparation method for the sensor is simple, high in selectivity, sensitivity and reproducibility, is good in stability and provides a good sensing platform for detecting the AFB1 in a practical sample.
Description
technical field [0001] The invention belongs to the technical field of biosensors, and in particular relates to a preparation method and application of a ratiometric electrochemical biosensor based on electrostatic interaction, which can be used for detecting mycotoxin aflatoxin B1 (AFB1). Background technique [0002] Mycotoxins are toxic secondary metabolites produced by certain fungi. Among all mycotoxins, AFB1 is the most toxic and poses a serious threat to human health (carcinogen, teratogen and mutagen), and has been listed as the first human carcinogen by the International Agency for Research on Cancer (IARC). AFB1 is easily produced in crops such as peanuts, corn, and wheat. Therefore, in order to protect the health of consumers and prevent food safety problems, the European Commission stipulates that the maximum content of AFB1 in grains is 2 μg / kg. At present, the existing detection methods of AFB1 have their own advantages and disadvantages. For example, high-pe...
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