Method for identifying breast cancer biomarkers and detection kit for breast cancer

A biomarker and breast cancer technology, applied in the field of medicine, can solve the problems of unsuitability for routine purposes, expensive instruments, cumbersome processing procedures, etc., and achieve the effects of simplified operation, reasonable instrument prices, and short analysis time

Inactive Publication Date: 2019-03-15
THE AFFILIATED HOSPITAL OF QINGDAO UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For example, when using enzymatic method, gas-liquid chromatography and high-resolution liquid chromatography, in order to avoid the influence of high-concentration glucose, it needs to be removed, resulting in cumbersome pretreatment process; gas-liquid chromatography-mass spectrometry instruments are expensive and not Suitable for routine purposes; large serum sample volume required
There is currently no literature reporting a pre-column 1-phenyl-5-methylpyrazolone (PMP)-derived HPLC method for the simultaneous detection of free mannose and glucose in breast cancer patient serum

Method used

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  • Method for identifying breast cancer biomarkers and detection kit for breast cancer
  • Method for identifying breast cancer biomarkers and detection kit for breast cancer
  • Method for identifying breast cancer biomarkers and detection kit for breast cancer

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0051] (1) Precisely weigh mannose (Man), glucosamine (GlcN), galactosamine (GalN), glucuronic acid (GlcUA), glucose (Glc), galactose (Gal), xylose (Xyl), rock Add appropriate amount of algalose (Fuc), add deionized water to prepare two mixed standard solutions containing the above monosaccharide 0.1mg / mL, and prepare immediately for use;

[0052] (2) Create an alkaline environment: Take 40 μL of the mixed monosaccharide standard in a 1.5mL EP tube, add 40 μL of 0.3mol / L sodium hydroxide, and vortex to mix;

[0053] (3) PMP derivatization: Add 60 μL 0.5mol / L 1-phenyl-5-methylpyrazolone (PMP) to each sample, vortex and mix well, and react in an oven at 70°C for 1 h after centrifugation;

[0054] (4) Neutralization reaction by adding acid: Take out the samples in the oven, let them cool down, add 40 μL of 0.3mol / L hydrochloric acid to each sample, and vortex to mix;

[0055] (5) Extraction: add 500 μL chloroform to each tube, vortex, centrifuge, remove the lower layer of chloro...

Embodiment 2

[0079] (1) Accurately weigh the appropriate amount of mannose (Man), rhamnose (Rha) and glucose (Glc), add deionized water to prepare 5 parts of the same mixed standard solution containing the above monosaccharide 0.1mg / mL, and use it immediately match;

[0080] (2) Create an alkaline environment: Take 40 μL of the mixed monosaccharide standard in a 1.5mL EP tube, add 40 μL of 0.3mol / L sodium hydroxide, and vortex to mix;

[0081] (3) PMP derivatization: Add 60 μL 0.5mol / L PMP to each sample, vortex and mix well, and react in an oven at 70°C for 1 h after centrifugation;

[0082] (4) Neutralization reaction by adding acid: Take out the samples in the oven, let them cool down, add 40 μL of 0.3mol / L hydrochloric acid to each sample, and vortex to mix;

[0083] (5) Extraction: add 500 μL chloroform to each tube, vortex, centrifuge, remove the lower layer of chloroform, keep the supernatant, repeat 3 times;

[0084] (6) Centrifuge the sample at 13000r / min for 10min, take 80μL su...

Embodiment 3

[0105] (1) Accurately weigh the appropriate amount of mannose and glucose, add deionized water to prepare the above monosaccharides containing 0.5mg / mL, 0.25mg / mL, 0.1mg / mL, 0.05mg / mL, 0.01mg / mL, 0.005mg / mL, 0.0025mg / mL, 0.001mg / mL, 0.0005mg / mL mixed standard solution, ready-to-use;

[0106] (2) Create an alkaline environment: Take 40 μL of the mixed monosaccharide standard in a 1.5mL EP tube, add 40 μL of 0.3mol / L sodium hydroxide, and vortex to mix;

[0107] (3) PMP derivatization: Add 60 μL 0.5mol / L PMP to each sample, vortex and mix well, and react in an oven at 70°C for 1 h after centrifugation;

[0108] (4) Neutralization reaction by adding acid: Take out the samples in the oven, let them cool down, add 40 μL of 0.3mol / L hydrochloric acid to each sample, and vortex to mix;

[0109] (5) Extraction: add 500 μL chloroform to each tube, vortex, centrifuge, remove the lower layer of chloroform, keep the supernatant, repeat 3 times;

[0110] (6) Centrifuge the sample at 130...

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Abstract

The invention provides a method for identifying breast cancer biomarkers and a detection kit for breast cancer. The biomarkers are free mannose and glucose obtained by high performance liquid chromatography derived from pre-column 1-phenyl-5-methyl pyrazolone (PMP) in serum. The detection method disclosed by the invention is a pre-column PMP derived high performance liquid chromatography. The technical scheme provided by the invention has the advantages of simple pretreatment, short analysis time, reasonable instrument price, accordance with conventional use, simple and easy-to-learn operationsteps and high accuracy of test results; normal people can be distinguished from breast cancer patients by only carrying out blood sampling and high performance liquid chromatography analysis; in addition, the required serum quantity is extremely low and the blood sampling amount is smaller than 1mL, and the like. An obtained result shows that the analysis method can quickly quantify the free mannose and glucose in the serum of breast cancer patients, and is of great significance for studying the relationship between the free mannose and glucose in the serum and breast cancer and finding a novel clinical detection marker for the breast cancer.

Description

technical field [0001] The invention belongs to the technical field of medicine, and in particular relates to a method for identifying biomarkers of breast cancer and a detection kit thereof. Background technique [0002] Breast cancer ranks first among female cancers. According to the 2012 Global Cancer Report (Globocan 2012) released by the International Cancer Research Institute (IARC), there are about 1.676 million new breast cancer cases and about 522,000 deaths among women worldwide every year. Currently, breast cancer screening methods commonly used in the world include clinical breast examination (CBE), mammography (MAM), ultrasound imaging (US) and magnetic resonance imaging (MRI). Wait. Although breast cancer mortality can be reduced, there are certain limitations. Currently the most commonly used clinical detection markers in the determination of tumor markers are CA125, CA19-9 and CEA. Blood examination is an ideal detection method because of its simple operati...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/02G01N30/06
CPCG01N30/02G01N30/06G01N2030/067
Inventor 张丽娟曾鹏娇李慧
Owner THE AFFILIATED HOSPITAL OF QINGDAO UNIV
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