Fluorescence ratio type immunoassay method for detecting fenitrothion

A phosphalt and immunoassay technology, applied in the field of immunoassay, can solve the problems of low signal-to-noise ratio, low precision, and the fluorescence signal is easily affected by external environmental factors, and achieves good interference ability, high precision, The effect of good stability

Active Publication Date: 2021-07-09
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, most of the current fluorescent immunoassay methods are based on the determination of a single fluorescence intensity "turn-off" or "turn-on". The fluorescence signal is easily affected by external environmental factors, resulting in low signal-to-noise ratio and low precision.
Moreover, there is no report on the fluorescent ratiometric immunoassay method for fenitrothion

Method used

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  • Fluorescence ratio type immunoassay method for detecting fenitrothion
  • Fluorescence ratio type immunoassay method for detecting fenitrothion
  • Fluorescence ratio type immunoassay method for detecting fenitrothion

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Embodiment 1

[0038] This embodiment provides a fluorescent ratiometric immunoassay method for detecting fenitrothion, which specifically includes the following steps:

[0039] Step S11. Synthesis of chitosan-modified platinum nanoparticles: Dissolve 0.1g chitosan in 50mL 1% HAc solution, draw 47mL chitosan solution in a beaker, add 2mL dropwise to the solution under stirring Concentration of 10mM H 2 PtCl 6 , stirred at room temperature for 30 min. Add 1 mL of freshly prepared NaBH with a concentration of 0.2M 4 solution and stirred at room temperature for 90 min to obtain Ch-Pt NPs.

[0040] From figure 1 , 2 It can be seen that the prepared chitosan platinum nanoparticles are round particles with a particle diameter of about 5 nm, and chitosan-modified platinum nanoparticles were successfully synthesized.

[0041] Step S12. Coating of microplates: pipette 100 μL of fenitrothion to coat the original microplates, put it in a 37°C incubator for coating for 8 hours, take out and wash t...

Embodiment 2

[0048] This embodiment provides a fluorescent ratiometric immunoassay method for detecting fenitrothion, which specifically includes the following steps:

[0049] Step S21. Synthesis of chitosan-modified platinum nanoparticles: Dissolve 0.2g chitosan in 50mL 1% HAc solution, draw 47mL chitosan solution in a beaker, add 2mL dropwise to the solution under stirring Concentration of 5mM H 2 PtCl 6 , stirred at room temperature for 50 min. Add 1 mL of freshly prepared NaBH with a concentration of 0.1M 4 solution and stirred at room temperature for 120 min to obtain Ch-Pt NPs.

[0050] Step S22. Coating of the microplate: absorb 100 μL of fenitrothion to coat the original microplate, put it in a 37°C incubator for coating for 10 hours, take out the coating solution after washing; add 120 μL of blocking solution to seal for 2 hours, shake off Dry the blocking solution and dry it at 37°C.

[0051] Step S23. Establishment of the standard curve: add 50 μL of fenitrothion standard s...

Embodiment 3

[0054] This embodiment provides a fluorescent ratiometric immunoassay method for detecting fenitrothion, which specifically includes the following steps:

[0055] Step S31. Synthesis of chitosan-modified platinum nanoparticles: Dissolve 0.5g chitosan in 50mL 2% HAc solution, draw 47mL chitosan solution in a beaker, add 4mL dropwise to the solution under stirring Concentration of 20mM H 2 PtCl 6 , stirred at room temperature for 60 min. Add 2 mL of freshly prepared NaBH with a concentration of 0.5M 4 solution and stirred at room temperature for 120 min to obtain Ch-Pt NPs.

[0056] Step S32. Coating the microplate: pipette 100 μL fenitrothion to coat the original microplate, put it in a 37° C. Dry the blocking solution and dry it at 37°C.

[0057] Step S33. Establishment of the standard curve: add 50 μL of fenitrothion standard solution of different concentrations and 50 μL of antibody solution into the microwell plate and mix, after incubation for 40 min, wash the plate w...

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Abstract

The invention provides a fluorescence ratio type immunoassay method of fenitrothion. A microporous plate is coated with a fenitrothion coating antigen, a fenitrothion nano antibody is fused with an alkaline phosphatase (Nb-ALP) recognition and signal amplification element, fenitrothion in a sample and the coating antigen on the plate compete to be combined with Nb-ALP, then redundant drugs and antibodies are washed away, and the detection result is accurate. the subsequently added L-ascorbic acid-2-trisodium phosphate (AAP) is catalyzed by the Nb-ALP connected to the microwell plate, and the L-ascorbic acid-2-trisodium phosphate (AAP) is hydrolyzed to generate ascorbic acid (AA); then, o-phenylenediamine (OPD) and chitosan modified platinum nanoparticles (Ch-Pt NPs) are added into the solution, and AA and OPD are competitively catalyzed and oxidized by the Ch-Pt NPs with oxidation-like characteristics. The OPD is oxidized into a fluorescent substance 2, 3-diaminophen (DAP), and the fluorescence emission peak is 568nm; AA in the oxidation state reacts with OPD to generate a quinoxaline derivative (DFQ), and the fluorescence emission peak is 430 nm; and therefore, the content of the fenitrothion in the sample is more sensitively reflected on the fluorescence ratio of 430nm to 568nm, and the purpose of high-sensitivity detection of the fenitrothion pesticide is achieved.

Description

technical field [0001] The invention belongs to the technical field of immunoassay, and more specifically relates to a fluorescence ratio immunoassay method for detecting fenitrothion. Background technique [0002] Pesticides refer to agents used in agriculture to control diseases and insect pests, regulate plant growth, and kill weeds. As a common organophosphorus pesticide, fenitrothion has been widely used as an insecticide in vegetable and fruit cultivation. However, the pesticide pollution caused by the widespread use and improper handling of fenitrothion has caused certain negative effects on the environment and public health. As a kind of organophosphorus pesticide, fenitrothion can inhibit the activity of acetylcholinesterase in organisms. Even if the concentration is relatively low, it will also inhibit choline and cause damage to central nervous system transmission, which will further damage human health; long-term low concentration accumulation may also cause ca...

Claims

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Application Information

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IPC IPC(8): G01N33/543G01N33/53G01N21/64
CPCG01N33/54313G01N33/54306G01N33/5308G01N21/6428G01N2430/10
Inventor 罗林吴卓裕徐振林王弘孙远明沈玉栋雷红涛
Owner SOUTH CHINA AGRI UNIV
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