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ELISA detection reagent kit suitable for diazepam relict analysis

An enzyme-linked immunosorbent assay and residue analysis technology, applied in analytical materials, measuring devices, instruments, etc., can solve the problems of late start of research, inability to meet the requirements of rapid, convenient and accurate detection, and achieve long storage time and pretreatment. Simple process and less time-consuming effect

Inactive Publication Date: 2008-12-03
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the detection of diazepam at home and abroad mainly includes high-performance liquid chromatography (HPLC), gas chromatography (GC), thin-layer chromatography (TLC), gas-mass spectrometry (GC-MS), liquid-mass However, these methods not only require expensive instruments and equipment, but also have relatively high requirements for test materials, which require further purification treatment, which can no longer meet the requirements of modern detection for fast, convenient and accurate
In recent years, research on immunoassay methods for benzodiazepines has been carried out abroad, but domestic research in this area started relatively late, and most of the detection products still need to rely on foreign imports

Method used

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  • ELISA detection reagent kit suitable for diazepam relict analysis
  • ELISA detection reagent kit suitable for diazepam relict analysis
  • ELISA detection reagent kit suitable for diazepam relict analysis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] Embodiment 1 Kit operation and result calculation:

[0018] After the samples to be tested were pretreated, they were made to volume with PBST for later use. Unpack the vacuum packaging bag and take out the microtiter plate, and equilibrate at room temperature for 5 minutes for later use. Prepare 0ng / mL, 0.7ng / mL, 1.4ng / mL, 2.8ng / mL, 5.6ng / mL, 11.2ng / mL, 56ng / mL, 112ng / mL diazepam standard solution, add 50μL standard sample or treat Put the good samples to be tested into each well, make 4 replicates of the standard sample and the sample, add 50 μL of diluted antibody, and incubate at 37°C for 30 minutes; pour out the liquid in the well, wash 5 times with diluted PBST, and remove the enzyme label. Invert the plate and pat it on absorbent paper; add 100 μL of enzyme-labeled goat anti-rabbit secondary antibody diluted at 1:3000, and incubate at 37°C for 30 minutes; pour out the liquid in the well, wash the plate 5 times with diluted PBST, and pat Dry; Mix liquid A and li...

Embodiment 2

[0020] Example 2 The formation of the enzyme-linked immunosorbent assay kit for diazepam residue analysis:

[0021] In this example, the kit contains the following parts:

[0022] (1) Enzyme label coated with diazepam antigen;

[0023] (2) Sponge bracket;

[0024] (3) 1mg / mL diazepam standard substance (Sigma company);

[0025] (4) Diazepam polyclonal antibody;

[0026] (5) horseradish peroxidase-labeled goat anti-rabbit antibody (Kangcheng Bioengineering Company);

[0027] (6) The formula of concentrated washing liquid is: 8g sodium chloride, 0.2g potassium dihydrogen phosphate, 3g disodium hydrogen phosphate, 0.4g potassium chloride, 0.5mL Tween-20 and 20mL distilled water;

[0028] (7) Formula of chromogenic solution A: 0.933g citric acid, 3.68g Na 2 HPO 4 12H 2 O, 18 μL 30% HO 2 o 2 and 100mL ultrapure water

[0029] (8) Chromogenic solution B formula: 60mg tetramethylbenzidine dissolved in 100mL ethylene glycol

Embodiment 3

[0030] Embodiment 3 shelf life experiment:

[0031] Store the kit at 4°C, take 0, 10, 20, 30, 60, 90, 120, 150 and 180d kits respectively, and use the antigen working concentration of 5.25 μg / mL and antibody working concentration of 1.2 μg / mL as the working concentration Concentration, carry out standard sample detection to determine its detection effect. The results of shelf life determination are shown in Table 1, indicating that IC 50 There is little change, and the kit can be stored at 4°C for more than 6 months.

[0032] Table 1 Kit preservation experiment

[0033]

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Abstract

The invention relates to an enzyme-linked immune detection kit suitable for the analysis of diazepam residues and belongs to the technology field of the enzyme-linked immune adsorption analysis kit. The kit comprises an enzyme label plate coated with the envelope antigen of the diazepam, a sponge support, a diazepam standard, a diazepam polyclonal antibody, an enzyme sign second antibody, a concentrating and washing liquid, a colored solution and a reaction stopping solution; the envelope antigen of the diazepam is the coupling compound of 3-half succinate diazepam and an egg-white protein; and the enzyme sign second antibody is a horseradish peroxidase labeled goat anti-rabbit antibody. The kit adopts the diazepam polyclonal antibody, and can accurately and sensitively detect the diazepam residues or other structurally similar benzodiazepine residues in urine or tissue; the process of pretreating samples is simple; the time consumption is low; a large quantity of samples can be tested simultaneously; furthermore, the cost for sample detection is lower than that of the traditional instrument detection method; the kit has a long retention time, no radioactive pollution, and practical significance for realizing on-site monitoring of the diazepam residues of the large quantity of samples.

Description

technical field [0001] The invention relates to an enzyme-linked immunoassay kit suitable for diazepam (DZP) residue analysis, belonging to the technical field of enzyme-linked immunosorbent assay (ELISA) kits. Background technique [0002] Diazepam is a benzodiazepine sedative and hypnotics, chemical name: 1-methyl-5-phenyl-7-chloro-1,3-dihydro-2H-1,4-benzodiazepine Zol-2-one. Used as an animal feed additive, as a growth promoter, it can make animals sleepy and less active, grow faster and change the meat quality. It is also used for sedation before slaughtering and animal quarantine, or before transferring animals to slaughter, so as to reduce animal tension and reduce animal injury and mortality. Drowsiness, weakness, headache, dizziness, nausea, constipation, and occasional rash, liver damage, bone marrow suppression and other adverse reactions can be seen with these drugs. If such drugs are randomly added to the feed, the accumulated drugs will enter the human body t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/543
Inventor 胥传来李秋生刘丽强袁缓彭池方胡拥明
Owner JIANGNAN UNIV
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