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Method for quantum dot mark indirect competition fluoroimmunoassay detection for becort

A technology of betamethasone and fluorescence immunity, which is applied in the field of immunoassay, can solve the problems of cumbersome operation and time-consuming, and achieve the effect of simple operation, strong fluorescence intensity and long fluorescence stability time

Inactive Publication Date: 2008-11-19
JIANGNAN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This type of method is to coat the enzyme-labeled plate with the original coating, add drugs and anti-drug antibodies, then add the enzyme-labeled secondary antibody, that is, the detection antibody, and finally add the substrate for color development. After a certain period of time, use a microplate reader to detect a certain Calculate the concentration of the drug to be tested in the sample based on the absorbance value of a specific wavelength based on the known content of the standard, which is cumbersome and time-consuming

Method used

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  • Method for quantum dot mark indirect competition fluoroimmunoassay detection for becort
  • Method for quantum dot mark indirect competition fluoroimmunoassay detection for becort

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] (1) Use denatured bovine serum albumin (dBSA) to wrap green QD590 labeled anti-betamethasone polyclonal antibody

[0029] (1) BSA degeneration:

[0030] Dissolve 16.5mg BSA in 10mL double-distilled water, add 0.42mg NaBH under stirring 4 ,Reaction at room temperature for 1h, heating at 60-80℃ for 20min to decompose excess NaBH 4 , BSA is denatured, the disulfide bond opens into -SH, and the final concentration of the dBSA aqueous solution is 5×10 -5 M.

[0031] (2) Denatured BSA wrapped quantum dots (dBSA-QDs):

[0032] The dBSA and quantum dots chromium dysprosium (CdTe) are mixed at a certain molar ratio (1:1), heated in a water bath at 60-80°C for 15 minutes, and kept at room temperature for two days to complete the package.

[0033] (3) Denatured BSA coated quantum dot conjugated antibody

[0034] Mix 5 μL 1-ethyl-(3-dimethylaminopropyl) carbodiimide EDC (0.056M) and 5 μL thio N-hydroxysuccinimide sulfo-NHS (0.1M) for 10 seconds, then add to 25μL dBSA-QDs(2×10 -5 In M),...

Embodiment 2

[0041] Example 2 Addition and recovery experiment

[0042] (1) Extraction and purification of the sample: Add 10mL of acetonitrile / water (7:3) mixed solution to 2g of chicken meat sample, vortex for 1min, ultrasonic for 30min, centrifuge at 2000×g for 10min, draw 2.5mL of supernatant liquid and clean Add 4 mL of n-hexane and 1 mL of dichloromethane to the glass tube for degreasing, vortex for 1 min to separate the three phases, draw 1 mL of the intermediate phase (corresponding to 0.2g sample) in a clean test tube; 50℃, slow N 2 Flow dry, and dissolve the residue with 0.2 mL of standard diluent (phosphate buffered saline PBST containing Tween-20 containing 10% methanol) and use it as a sample for analysis.

[0043] (2) Add betamethasone standard solution to 2g blank chicken sample to make the concentration of 1ng / g, 5ng / g, 10ng / g, 20ng / g, and prepare five samples for each concentration. The processing method is analyzed after extraction. The results are shown in Table 1. It can be...

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Abstract

Disclosed is a method of quantum dot-labeled indirect competitive fluorescence immunoassay of betamethasone, which belongs to the immunoassay method technique field. Quantum dots for labeling antibodies of the invention have the emission spectra of QD590, and the method comprises: directly covering coating antigens in micro-holes of an enzyme label plate, adding betamethasone standard solution or a sample under test to form an antigen-antibody fluorescence immunity compound body, stimulating and detecting the fluorescence intensity of the formed antigen-antibody fluorescence immunity compound body with a fluorescence enzyme-labeling instrument, and obtaining the concentration of betamethasone in the sample under test through comparing with the standard solution. The invention can detect the content of betamethasone in the sample under test without adding chromogenic substance, namely, the concentration of betamethasone in the sample under test can be detected indirectly through the fluorescence intensity of the antigen-antibody immunity compound body, and both the operation and reaction need only one step; and the quantum dots for labeling antibodies of the invention have advantages of stronger emitted fluorescence intensity and long stabilization time of fluorescence compared with the traditional fluorescence.

Description

Technical field [0001] A quantum dot-labeled indirect competitive fluorescence immunoassay method for detecting betamethasone belongs to the technical field of immunoassay methods. Background technique [0002] Betamethasone (Betamethasone, BET) is a synthetic glucocorticoid, the chemical name is 16α-methyl-11β, 17α, 21-trihydroxy-9α-fluoropregesta-1, 4-diene-3, 20 two ketone. It has the effects of anti-allergic, anti-inflammatory and influencing glucose metabolism. It is often used to treat inflammatory reactions, immune diseases, cattle ketosis and sheep pregnancy toxemia in livestock. Dexamethasone is also often used as a growth promoter to increase the feed intake of livestock to achieve weight gain. However, the toxicology test proved that the drug has mutagenicity and cumulative toxicity, with an ADI value of 0.000015mg / kgbw / day. If the drug is added to the feed at will, the accumulated drug enters the human body through the food chain, which will cause great harm. For this...

Claims

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Application Information

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IPC IPC(8): G01N33/74G01N33/543G01N21/64
Inventor 胥传来袁媛陈伟彭池方谢会玲郝晓蕾
Owner JIANGNAN UNIV
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