Aptamer biosensor preparation method adopting ferrocene as electron donor, and application of aptamer biosensor

A biosensor and electron donor technology, applied in the field of electrochemical biosensor detection, can solve the problems of expensive equipment, time-consuming and laborious, and high technical requirements for operators, and achieve the effect of simple preparation method, high stability and specificity

Active Publication Date: 2019-11-29
SHANDONG UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, most of them are time-consuming and labor-intensive, the instruments and equipment are expensive, and the operators have high technical requirements, so they are not suitable for rapid on-site analysis.

Method used

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  • Aptamer biosensor preparation method adopting ferrocene as electron donor, and application of aptamer biosensor
  • Aptamer biosensor preparation method adopting ferrocene as electron donor, and application of aptamer biosensor
  • Aptamer biosensor preparation method adopting ferrocene as electron donor, and application of aptamer biosensor

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] A method for preparing an aptamer biosensor based on ferrocene as an electron donor, the specific steps of the preparation method are:

[0026] (1) Design of the hairpin aptamer (HP) and preparation of its solution

[0027] At the 3′ end of the aptamer (5′-AGCTTGCTGCAGCGATTCTTGATCGCCACAGAGCT-3′), the base pair CAAGCT complementary to the first few base pairs (AGCTTG) at the 5′ end is added, and amino groups (-NH 2 ) and ferrocene (Fc), a newly synthesized aptamer chain (NH 2 -AGCTTGCTGC AGCGATTCTTGATCGCCACAGAGCTCAAGCT-Fc) was fitted and analyzed with DNAman software, and it was found that the hairpin structure could be effectively formed. In order to make the designed and synthesized aptamer form a hairpin structure, the aptamer stock solution was heated in a water bath at 95°C for 5 min, and then slowly cooled to room temperature. It was diluted to a 2 μM stock solution with Tris-HCl at pH 7.5 as solvent.

[0028] (2) Preparation of graphene oxide-chitosan (GO-CS) s...

Embodiment 2

[0039] Application of an aptamer biosensor based on ferrocene as an electron donor:

[0040] (1) Detection of organophosphorus pesticides in water

[0041] 8 μL of different organophosphorus pesticides at different concentrations were dropped onto the aptamer biosensor and incubated in air for 1 h. After being gently rinsed with ultrapure water and blown dry with nitrogen, the DPV electrochemical signals were measured in pH 7.4 phosphate buffer solution. The result is as Figure 4-7 shown ( Figure 4 profenofos, Figure 5 Phorate, Figure 6 isocarbophos, Figure 7 Omethoate), there is a good linear relationship between the DPV response signal and the logarithm of the concentration of the four organophosphorus pesticides. Their linear regression equations, linear ranges, correlation coefficients and detection limits are summarized in Figure 8 middle. The extremely low detection line of the sensor shows that the aptamer biosensor prepared by the present invention ha...

Embodiment 3

[0045] Performance test of an aptamer biosensor based on ferrocene as an electron donor:

[0046] (1) Specificity analysis

[0047] The selectivity of the aptasensor was tested using carbaryl, malathion, methamidophos, monocrotophos, chlorpyrifos and their mixtures as interfering pesticides. Such as Figure 10 Shown (a, b, c, d, e, f represent five interfering pesticides and their mixtures; g, h, i, j, k represent profenofos, phorate, isocarbophos and oxo Dimethoate and its mixture; l represents the mixture of these nine pesticides; all pesticides are at 10 nM), and when there are no specific targets (profenofos, phorate, isocarbophos and omethoate), the Larger chemical signals indicate that the aptamer cannot bind to these interfering pesticide molecules. In addition, there is no obvious difference between histogram k and l, which once again proves that the aptamer biosensor prepared by the present invention has good specificity and anti-interference ability.

[0048] (2)...

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Abstract

The invention relates to an aptamer biosensor preparation method adopting ferrocene as an electron donor. According to the method, a chitosan solution is added into a carboxyl-activated graphene oxidesolution, and the surface of a screen-printed carbon electrode is modified with an obtained mixed solution; a specific base pair is added to the tail end of an aptamer, so that a hairpin structure can be designed and synthesized, two ends of the aptamer are modified with amino and ferrocene respectively, and the aptamer chain is effectively fixed to the surface of the graphene oxide-chitosan / silk-screen printed carbon electrode through the formation of amido bonds; the specific recognition of the aptamer and a target object induces the conformation change of the aptamer, so that current signals generated by the ferrocene in a phosphate buffer solution is changed, so that the target object can be monitored. According to the invention, the ultrasensitive detection of four organophosphorus pesticides including profenofos, phorate, isocarbophos and omethoate can be realized; detection time is short; detection limit is low; and a new technology is provided for the detection of organophosphorus pesticides and harmful substances in other fields.

Description

technical field [0001] The invention relates to a preparation method and application of an aptamer biosensor based on ferrocene as an electron donor, and belongs to the technical field of electrochemical biosensor detection. Background technique [0002] Organophosphorus pesticides are phosphorus-containing organic compounds that are widely used in agriculture to control pests and weeds because of their high efficiency, low toxicity, and low residue. The use of these pesticides can help prevent crop loss. However, excessive organophosphorus pesticides remaining on agricultural products can enter the human body through the food chain, combine with acetylcholinesterase to cause neurological disorders, lead to paralysis or even death, and cause great harm to human health. Profenofos, phorate, isocarbophos and omethoate are widely used organophosphorus pesticides, and incidents of exceeding standards often occur in vegetable products. Therefore, it is necessary to construct a ...

Claims

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Application Information

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IPC IPC(8): G01N27/327G01N27/48
CPCG01N27/3275G01N27/3277G01N27/48
Inventor 郭业民符家韵史孝杰孙霞
Owner SHANDONG UNIV OF TECH
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