Gentamycin immunoassay reagent as well as preparation method and detection method thereof

A technology for gentamicin and immunological detection, applied in biological testing, measuring devices, material inspection products, etc., can solve the problems of low measurement accuracy and complicated operation of gentamicin detection process, and achieve high sensitivity and specificity Strong, Quantitatively Accurate Effects

Inactive Publication Date: 2018-06-01
太原瑞盛生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The purpose of the present invention is to solve the problems of complex operation and low measurement accuracy in the detection process of gentamicin in the prior art. Gentamicin homogeneous enzyme immunoassay reagent and preparation method thereof

Method used

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  • Gentamycin immunoassay reagent as well as preparation method and detection method thereof
  • Gentamycin immunoassay reagent as well as preparation method and detection method thereof
  • Gentamycin immunoassay reagent as well as preparation method and detection method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Example 1: Preparation of glucose dehydrogenase-antigen conjugate

[0044] 1) Preparation of glucose dehydrogenase (GDH) solution:

[0045] a. Weigh 10 mg of GDH with a specification of 100KU, dissolve it in 8 ml of PBS solution with a concentration of 10mM at room temperature, pH=7.4;

[0046] 2) Activation of gentamicin

[0047] Add the following chemicals to a beaker and stir to dissolve: 25 mg gentamicin, 20 mg 1-ethyl-3-(-3-dimethylaminopropyl)carbodiimide, 5 mg N-hydroxysulfosuccinyl Imine, dissolved in 600 μL morpholineethanesulfonic acid (MES) solution, stirred and dissolved at room temperature, and reacted for 30 minutes;

[0048] 3) Coupling of GDH and gentamicin

[0049] a. the above-mentioned activated gentamicin solution is added dropwise in the above-mentioned dissolved GDH solution;

[0050] b Stir overnight at 2-8°C;

[0051] 4) Purify the conjugated enzyme-labeled antigen

[0052] The conjugated enzyme-labeled antigen was purified by G-25 gel chro...

Embodiment 2

[0053] Example 2: Preparation of Gentamicin Homogeneous Enzyme Immunoassay Reagent

[0054] Gentamicin homogeneous enzyme immunoassay reagents, including: enzyme-labeled gentamicin, indicator reagent for detecting gentamicin antibody-enzyme-labeled gentamicin complex. The indicator reagent is selected from enzyme reagents, radioisotope reagents, fluorescent reagents and chemiluminescent reagents. Preferably, the indicator reagent is an enzyme reagent, including: an enzyme-labeled conjugate and an enzyme substrate. Wherein, the enzyme-labeled conjugate includes glucose dehydrogenase-antigen conjugate, which can be obtained by chemical synthesis.

[0055]Before using the gentamicin homogeneous enzyme immunoassay reagent, in order to avoid the reaction between the enzyme-labeled conjugate and the enzyme substrate in the indicator reagent, the enzyme-labeled conjugate and the enzyme substrate are placed separately, so Qing Ampicillin Homogeneous Enzyme Immunoassay Reagent consis...

Embodiment 3

[0064] Example 3: Gentamicin Homogeneous Enzyme Immunoassay Reagent Reaction Calibration Curve.

[0065] 1) Preparation of calibrator: Dissolve commercially available human gentamycin recombinant protein in a solution similar to human serum matrix (NaCl0.9%, BSA0.2%, NaN 3 0.1%, Tris-HCl pH 7.4), and made different concentrations of calibrators. Taking the gentamicin calibrator of Suzhou Jianglai Institute of Biology as the original standard, the gentamicin kit was used to test the calibrator of different concentrations 10 times, and the average value was obtained to obtain the concentration of the gentamicin calibrator : 1, 5, 10, 20, 40, 80 ng / mL.

[0066] 2) Detection of biochemical analyzer: take the operation of Hitachi 7170 as an example: the measurement wavelength is 340nm, take different concentrations of calibrator solutions (15μl), add gentamicin R 1 Reagent (160 μl), mix well, then add Gentamicin R 2 Reagent (40μl), after mixing, measure the OD at different time ...

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Abstract

The invention discloses a gentamycin immunoassay reagent as well as a preparation method and a detection method thereof. The gentamycin immunoassay reagent comprises enzyme labeled gentamycin and an indicator reagent used for detecting a gentamycin antibody-enzyme labeled gentamycin compound; and the enzyme labeled gentamycin formed by coupling the gentamycin to glucose dehydrogenase. The gentamycin immunoassay reagent provided by the invention can accurately and rapidly determine the content of the gentamycin in a sample such as human blood. Compared with the existing detection reagent on themarket, the gentamycin immunoassay reagent has the advantages of convenience, rapidness, high sensitivity, strong specificity, accuracy in quantification and the like and is beneficial to clinical popularization and use.

Description

technical field [0001] The invention relates to the field of biological detection, in particular to a gentamicin immunodetection reagent and a preparation and detection method thereof. Background technique [0002] Gentamicin (GM) is an aminoglycoside antibiotic with a broad antibacterial spectrum. Its mechanism of action is to act on ribosomes in bacteria, inhibit bacterial protein synthesis, and destroy the integrity of bacterial cell membranes. GM is widely used in the pest control of agriculture, livestock, poultry, forestry and aquatic products. Due to the abuse of users, there is often a problem of large GM residues. Moreover, gentamicin has obvious toxic effects on humans, and has serious damage to cranial nerves, hearing and kidneys. Therefore, it is very necessary to limit and detect the residues of gentamicin in food of animal origin. [0003] Currently, the commonly used methods for detecting gentamicin include high performance liquid chromatography, enzyme-lin...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/535G01N33/53
CPCG01N33/535G01N33/5308G01N2430/00
Inventor 张轩马运乐杨钰祥于雪纯杜爱铭徐兵
Owner 太原瑞盛生物科技有限公司
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