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Adiponectin receptor ligand screening method and adiponectin receptor agonist screening method

A screening method, adiponectin technology, applied in material excitation analysis, material test, fluorescence/phosphorescence, etc., can solve the problems of limited sample processing, long experiment time, low effectiveness, etc., to achieve high accuracy and easy operation. Simple, low-cost effect

Inactive Publication Date: 2013-01-16
CHENGDU MEDICAL COLLEGE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, these techniques have many limitations, for example, the samples that can be processed at one time are extremely limited (up to 8-12 samples per gel), and the experiment time of this technique is long, from the beginning of the experiment to the data (8 -12 samples) takes at least a week and, in most cases, can only prove again that these samples are "ligands", high cost and low effectiveness

Method used

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  • Adiponectin receptor ligand screening method and adiponectin receptor agonist screening method
  • Adiponectin receptor ligand screening method and adiponectin receptor agonist screening method
  • Adiponectin receptor ligand screening method and adiponectin receptor agonist screening method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1 Screening of Adiponectin Receptor Ligands

[0043] A mixture of FITC-gAPN at a final concentration of 600 nM and AdipoR1 at a final concentration of 1 μM was prepared, and then added to a 384-well plate at 19 μL / well. The unknown compound was dissolved in 20 μL DMSO to prepare a 10 mM stock solution, and the stock solution was half-diluted 11 times to obtain 12 different concentrations of samples to be tested (the concentration range was 10 mM-4.88 mM). Add 1 μL / well of the sample to be tested into a 384-well plate (final concentration range 500 μM-0.25 μM). Shake the 384-well plate for 30 seconds before the test to mix the solution thoroughly, and use SpectraMax (M5) to read the polarized fluorescence value of the sample. The instrument should be set at Ex 554 / Em 575. Each concentration of each sample should be tested 3 times.

[0044] After the instrument reads the polarized fluorescence value, the obtained data is transferred to SigmaPlot 11 for data pro...

Embodiment 2

[0045] Example 2 Screening of Adiponectin Receptor Ligands

[0046] A mixture of FITC-gAPN at a final concentration of 600nM and AdipoR2 at a final concentration of 1 μM was prepared, and then 19 μL / well of this mixture was added to a 384-well plate (final concentration range 500 μM-0.25 μM). The unknown compound was dissolved in 20 μL DMSO to prepare a 5 mM stock solution, which was half-diluted 11 times to obtain 12 different concentrations of samples to be tested (the concentration range was 10 mM-4.88 mM). Add 1 μL / well of the sample to be tested into a 384-well plate. Shake the 384-well plate for 30 seconds before the test to mix the solution thoroughly, and use SpectraMax (M5) to read the polarized fluorescence value of the sample. The instrument should be set at Ex 554 / Em 575. Each concentration of each sample should be tested 3 times.

[0047] After the instrument reads the polarized fluorescence value, the obtained data is transferred to SigmaPlot 11 for data proce...

Embodiment 3

[0048] Example The screening of triadiponectin receptor ligand

[0049] A mixture of FITC-gAPN at a final concentration of 100 nM and AdipoR1 at a final concentration of 1 μM was prepared, and then added to a 384-well plate at 19 μL / well. The unknown compound was dissolved in 20 μL DMSO to prepare a 10 mM stock solution, and the stock solution was diluted 7 times in half to obtain 8 different concentrations of the samples to be tested (the concentration range was 10 mM-78 nM). Add 1 μL / well of the sample to be tested into a 384-well plate (final concentration range 500 μM-4 μM). Shake the 384-well plate for 30 seconds before the test to mix the solution thoroughly, and use SpectraMax (M5) to read the polarized fluorescence value of the sample. The instrument should be set at Ex 554 / Em 575. Each concentration of each sample should be tested 3 times.

[0050] After the instrument reads the polarized fluorescence value, the obtained data is transferred to SigmaPlot 11 for data...

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Abstract

The invention discloses an adiponectin receptor ligand screening method which uses the polarized fluorescence technology for screening adiponectin receptor ligands. The invention further discloses an adiponectin receptor agonist screening method including the steps: taking a to-be-detected sample, adopting the adiponectin receptor ligand screening method for screening, and screening by the aid of the cell culture immune polarized fluorescence technology. The adiponectin receptor ligand screening method and the adiponectin receptor agonist screening method can accurately and effectively screen the adiponectin receptor ligands and adiponectin receptor agonists, lay a good foundation for screening anti-inflammation, antidiabetic and atherosclerotic drugs and have bright industrial application prospect.

Description

technical field [0001] The invention relates to a method for screening adiponectin receptor ligands, belonging to the field of biotechnology. Background technique [0002] Adipose tissue is mainly composed of a large number of adipocytes aggregated into groups. Adiponectin is an endogenous bioactive polypeptide or protein secreted by adipocytes. There are 6 forms of existence in the body: gAPN, fAPN , LMW, MMW, HMW and Alb-LMW. Among them, fAPN is the basic unit of other various forms, while LMW, MMW, and HMW are oligomers of fAPN, and gAPN is the spherical part of fAPN. Adiponectin is an insulin-sensitizing hormone (AnInsulin-sensitizing Hormone), which can improve insulin resistance (Insulin resistance) and arteriosclerosis in mice; studies on humans have found that adiponectin levels can predict type II diabetes and development of coronary heart disease, and demonstrated antidiabetic, antiatherogenic and inflammatory potential in clinical trials. [0003] Adiponectin h...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N21/64
Inventor 孙毅毅钟玲林东陈修平
Owner CHENGDU MEDICAL COLLEGE
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