Method and detection kit for identifying ovarian cancer biomarkers

A biomarker, ovarian cancer technology, applied in the field of medicine, can solve the problems of being unsuitable for routine purposes, cumbersome processing, large volume of serum samples, etc., and achieve the effects of simplified operation, short analysis time and simple operation steps

Inactive Publication Date: 2019-03-01
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 the serum of ovarian cancer patients

Method used

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

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

[0080] (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;

[0081] (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;

[0082] (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 hour after centrifugation;

[0083] (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;

[0084] (5) Extraction: add 500 μL chloroform to each tube, vortex, centrifuge, remove the lower layer of chloroform, keep the supernatant, repeat 3 times;

[0085] (6) Centrifuge the sample at 13000r / min for 10min, take 80μL...

Embodiment 3

[0106] (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;

[0107] (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;

[0108] (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 hour after centrifugation;

[0109] (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;

[0110] (5) Extraction: add 500 μL chloroform to each tube, vortex, centrifuge, remove the lower layer of chloroform, keep the supernatant, repeat 3 times;

[0111] (6) Centrifuge the sample at ...

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Abstract

The invention provides a method and a detection kit for identifying ovarian cancer biomarkers. The biomarkers are free glucose and free mannose which are obtained by subjecting serum to pre-column 3-methyl-1-phenyl-2-pyrazolin-5-one (PMP) derivatization high-performance liquid chromatography. A detection method is a pre-column PMP derivatization high-performance liquid chromatography. According tothe technical scheme, the method and the detection kit for identifying the ovarian cancer biomarkers have the advantages that the pre-treatment is simple, analysis time is short, the instrument priceis reasonable, the conventional use is met, the operation steps are simple and easy to learn, the detection result is high in accuracy, a normal person and an ovarian cancer patient can be distinguished by just sampling blood and subjecting the blood to the high-performance liquid chromatography for analysis, moreover, the required serum amount is very few, the amount of sampled blood is less than 1 mL, and the like. An obtained result shows that by means of the analysis method, the free mannose and glucose in the serum of the ovarian cancer patient can be fast quantified, so that the methodand the detection kit for identifying the ovarian cancer biomarkers have significant meaning to studying a relation between the free mannose and the free glucose in the serum and ovarian cancer and finding a novel ovarian cancer clinical detection marker.

Description

technical field [0001] The invention belongs to the technical field of medicine, and in particular relates to a method for identifying a biomarker of ovarian cancer and a detection kit thereof. Background technique [0002] Ovarian cancer is one of the most common tumors in female reproductive organs, ranking third in incidence after cervical cancer and uterine body cancer. However, the number of deaths due to ovarian cancer ranks first in all types of gynecological tumors, posing a serious threat to women's lives. The preoperative diagnostic examination methods for ovarian cancer generally include ultrasound, laparoscopic CT scan, magnetic resonance imaging, biopsy and other techniques, but none of them can give diagnostic results. CA125, the most commonly used clinical marker in the determination of tumor markers, is an ideal marker for ovarian epithelial carcinoma, and its positive detection rate can reach 70% to 90% in serous carcinoma. About 50% of patients with ovari...

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 张丽娟曾鹏娇李鹃王沛涛
Owner THE AFFILIATED HOSPITAL OF QINGDAO UNIV
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