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Homogeneous chemiluminescence immunoassay method for measuring for organophosphorus pesticide Dursban

A homogeneous chemiluminescence, organophosphorus pesticide technology, applied in the field of pesticide residue analysis and chemiluminescence immunoassay, can solve the problem of less pesticides and veterinary drugs, and achieve the effect of increasing sensitivity

Inactive Publication Date: 2011-05-11
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Chemiluminescent Immunoassay (CLIA) is a highly sensitive and specific analytical technique, which is widely used in bioengineering and medical clinical testing, but there are relatively few studies on agricultural and veterinary drugs at home and abroad.

Method used

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  • Homogeneous chemiluminescence immunoassay method for measuring for organophosphorus pesticide Dursban
  • Homogeneous chemiluminescence immunoassay method for measuring for organophosphorus pesticide Dursban
  • Homogeneous chemiluminescence immunoassay method for measuring for organophosphorus pesticide Dursban

Examples

Experimental program
Comparison scheme
Effect test

experiment example 2

[0045] 2. Experimental Example 2 Synthesis of Luminol-Bovine Serum Albumin (Lu-BSA)

[0046] Such as image 3 , Add 50mg of luminol to 5.0mL of 3.0mol / L HCl, stir on a magnetic stirrer at 0-4°C until dissolved. Add 50mg of NaNO to the acidic luminol solution at 0~4℃ 2 solid, and reacted in the dark for 15 minutes. When the reaction was completed, the mixed solution was blue to Congo red test paper, and blue to potassium iodide starch test paper, and then adjusted the mixed solution with 0.50ml / L pH of 8.5 borax buffer to make its pH as Around 8.5. Then add a certain amount of 0.10mol / L PBS buffer solution containing BSA, and react in the dark for 24 hours at 0-4°C. The reaction mixture was dialyzed by static gradient dialysis with borax buffer solution in a dark place, and the molecular weight cut-off of the dialysis membrane was (8000-12000). Take the dialysate every 12 hours, scan through the fluorescence spectrum, and detect the luminol (Lu) content of the dialysate wit...

experiment example 3

[0048] 3. Experimental example 3 Synthesis of chlorpyrifos chemiluminescent marker (Lu-BSA-AR)

[0049] Such as Figure 4, Weigh the activated chlorpyrifos (AR) and dissolve it in N,N-dimethylformamide (DMF), add N-hydroxysuccinimide, stir on a magnetic stirrer for 1h, then add 1-ethyl- (3-Dimethylaminopropyl) carbodiimide hydrochloride, continue to stir and react for 2h. A certain amount of the synthesized Lu-BSA solution was added to the above mixture, and the reaction was carried out under magnetic stirring at room temperature in a dark place for 5 hours. After the reaction finishes, the reaction mixture is dialyzed with phosphate buffered saline, and the dialysate is taken every 12 hours, and the content of activated chlorpyrifos (AR) in the dialysate is detected by ultraviolet spectrophotometry, and no AR can be detected in the dialysate after dialysis. Luminol-BSA-chlorpyrifos complex (Lu-BSA-AR) solutions are available. The chlorpyrifos chemiluminescent marker was pr...

Embodiment 7

[0056] 7. Drawing of luminescence kinetic curve of chlorpyrifos chemiluminescence marker (Lu-BSA-AR) in embodiment 7

[0057] Such as Figure 7 , respectively take a certain amount of Lu, Lu-BSA, Lu-BSA-AR solution, and use pH 9.6 carbonate buffer solution to make up to 10ml, then take 100μL of the solution into the measuring cup, inject 0.2mol / L of H 2 o 2 The solution was detected on a BPCL weak luminescence measuring instrument, the chemiluminescence intensity was recorded, and the luminescence kinetic curves of Lu, Lu-BSA, and Lu-BSA-AR were drawn. The results showed that the luminous intensity of Lu-BSA-AR and Lu-BSA decreased compared with Lu, while the luminous intensity of Lu-BSA-AR and Lu-BSA was similar, although the luminous intensity of Lu-BSA-AR was lower than that of Lu , but Lu-BSA-AR still showed good chemiluminescence kinetics.

[0058] 8. Preparation of Experimental Example 8 Working Curve

[0059] Such as Figure 9 , to fix the amount of chlorpyrifos ...

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Abstract

The invention discloses a homogeneous chemiluminescence immunoassay method for measuring an organophosphorus pesticide Dursban and belongs to the technical fields of pesticide residue analysis and chemiluminescence immunoassay. The method comprises the following steps: 1) activating the organophosphorus pesticide Dursban and introducing an active group carboxyl group so as to acquire the activated organophosphorus pesticide Dursban; 2) marking the chemiluminescence agent luminol onto bovine serum albumin (BSA) so as to acquire the luminol-BSA, wherein the coupling ratio reaches (1:20) to (1:40); 3) marking the activated organophosphorus pesticide Dursban onto the luminol-BSA, so as to acquire a Dursban chemiluminescence marker; 4) drawing a luminescence kinetics curve of the Dursban chemiluminescence marker, wherein a time axis is used as the horizontal ordinate and the relative luminous intensity is used as the vertical coordinate; and 5) establishing a homogeneous chemiluminescence immunoassay direct competition method to confirm the linear relationship between the luminous intensity and the antigen concentration of a sample to be identified. The method provided by the invention is suitable for the speedy qualitative and quantitative analysis detection for the Dursban on site.

Description

technical field [0001] The invention belongs to the technical fields of pesticide residue analysis and chemiluminescence immunoassay, and in particular relates to a homogeneous chemiluminescence immunoassay method for detecting organophosphorus pesticide chlorpyrifos. Background technique [0002] Chlorpyrifos (chlorpyrifos), O,O-diethyl-O-(3,5,6-trichloro-2-pyridyl)phosphorothioate. A high-efficiency insecticide popularized and used by Dow Chemical Company in 1965. It is used in the pest control of food, vegetables, fruits and commercial crops worldwide. It is one of the main varieties currently replacing highly toxic organophosphorus insecticides. It is also an important variety for the monitoring of pesticide residues in safe agricultural products, and the residue problems caused by crops such as grains have been reported. Moreover, the situation of chlorpyrifos residues found in environmental samples is becoming more and more serious, which poses a potential threat to p...

Claims

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Application Information

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IPC IPC(8): G01N33/566G01J1/00
Inventor 刘晓宇姚鑫石旺荣赵冬冬孟思王国磊刘荣飞
Owner HUAZHONG AGRI UNIV
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