CN102645540A - Blood marker for diagnosing hepatic fibrosis and early cirrhosis - Google Patents
Blood marker for diagnosing hepatic fibrosis and early cirrhosis Download PDFInfo
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- CN102645540A CN102645540A CN2012100947233A CN201210094723A CN102645540A CN 102645540 A CN102645540 A CN 102645540A CN 2012100947233 A CN2012100947233 A CN 2012100947233A CN 201210094723 A CN201210094723 A CN 201210094723A CN 102645540 A CN102645540 A CN 102645540A
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- blood
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- hepatic fibrosis
- galactose agglutinin
- liver fibrosis
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Abstract
The invention discloses a blood marker for diagnosing hepatic fibrosis and early cirrhosis. The blood marker refers to galectin 3. The concentration of the galectin 3 in the blood is analyzed by using the blood marker through enzyme linked immune reaction so as to analyze and evaluate hepatic fibrosis and early cirrhosis. By using the blood as the detection sample, early diagnosis on hepatic fibrosis and early cirrhosis can be achieved conveniently and rapidly.
Description
Technical field
The present invention relates to a kind of blood mark galactose agglutinin 3 that is used for diagnosing liver fibrosis and early-phase hepatocirrhosis, belong to biomedical sector.
Background technology
Liver fibrosis is to comprise virus hepatitis, alcoholic hepatitis, fatty liver, the common pathologic process that the liver tissue injury of different reasons such as drug induced hepatic injury changes to cirrhosis.Cirrhosis is irreversible liver tissue lesions, but does not develop into as yet in the process of cirrhosis in liver fibrosis, carries out pharmaceutical intervention timely, can realize that the inhibition of fibrotic processes and hepatic tissue are normal reparations, avoids the generation of cirrhosis.Yet, to the diagnosis of liver fibrosis, still need rely on the hepatic tissue aspiration biopsy at present, this process is traumatic, might cause fatefulue complication.Simultaneously; It is incompatible that early stage liver fibrosis does not have tangible physiology, and it is unpractical to judge Fibrotic that patient is carried out periodic hepatic tissue aspiration biopsy, therefore is badly in need of novel clinically; With blood is the mode that detects sample, with the convenient early diagnosis that liver fibrosis is taken place.
Summary of the invention
The purpose of this invention is to provide a kind of blood mark galactose agglutinin 3 that is used for diagnosing liver fibrosis and early-phase hepatocirrhosis.
Galactose agglutinin 3 described in the present invention, English name galectin-3, the detection of galactose agglutinin 3 can connect immunological technique and detects in the blood through the enzyme of maturation.Early-stage Study of the present invention is found, the stellate cells of the abnormal activation that plays a crucial role in the liver fibrosis generating process, and great expression galactose agglutinin 3, its expression is relevant with the activation degree of stellate cells.Simultaneously, we are also noted that in the serum of liver fibrosis mouse model, the content of galactose agglutinin significantly raises.
For detecting the galactose agglutinin 3 in the serum sample, the invention provides a kind of experimental technique that detects galactose agglutinin 3 in the blood sample through ELISA.
The experimental technique of above-mentioned ELISA is following: the antibody sandwich that will resist galactose agglutinin 3 is in ELISA Plate, and sample adds ELISA Plate, hatch certain hour after; Fully wash plate, add the antibody of the galactose agglutinin 3 of Avidin mark, hatch certain hour after; Fully wash plate, add biotin labeled horseradish peroxidase then, hatch certain hour after; Fully wash plate, add the reaction substrate of horseradish peroxidase at last, add the hydrochloric acid solution cessation reaction behind the reaction certain hour; Through the light absorption in the ELISA Plate hole after the ELIASA detection reaction, and compare, confirm the content of galactose agglutinin in the sample with standard items.
The present invention carries out the check and analysis assessment of liver fibrosis and early-phase hepatocirrhosis through the concentration of the galactose agglutinin in the enzyme linked immunoassay analyzing blood 3.
The collection processing method of blood sample is including, but not limited to following step: collect blood sample through the negative pressure anticoagulant tube, and through 500g centrifugal 5 minutes, remove haemocyte, obtain blood plasma, be sample to be tested.
The present invention is through carrying out illustration to the mouse experiment The Animal Model Study.The animal here includes, but are not limited to: mouse, rat, performing animal for example but be not limited to cat, dog, and some other animal for example but be not limited to ox, sheep, pig, horse and primate for example still are not limited to monkey and people.The interior detection of the body of hepatic fibrosis in mice empirical model for example still is not limited to the active model that detects of people's drug disposition as a reference by extensive recognition and acceptance to other biological.
Embodiment
Embodiment 1
The preparation of mouse carbon tetrachloride liver fibrosis mouse model:
Around getting age the ICR mouse, lumbar injection is dissolved in palmitic phenixin, dosage is the 1mg/kg body weight, weekly twice, continuously around, can obtain the animal model of remarkable liver tissue fibrosis.
Control animals injection equal volume, same number, the palm oil of identical time.
After model preparation finishes, the mode of getting blood through eyeball, in first week of modeling, second week, the 3rd week, obtain blood sample around the, and further through centrifugal acquisition plasma sample, for use.
Embodiment 2
Through the galactose agglutinin 3 in the enzyme linked immunoassay test experience model plasma sample:
1. encapsulate: encapsulating damping fluid with 0.05M PH9.0 carbonate, antibody (available from U.S. Abcam company) is diluted to protein content is 1～10 μ g/ml.In the reacting hole of each XPS, add 0.1ml, 4 ℃ are spent the night.Discard solution in the hole next day, washes 3 times each 3 minutes with lavation buffer solution.(being called for short washing, down together).
2. application of sample: the sample 0.1ml to be checked that adds certain dilution puts 37 ℃ and hatched 1 hour in the above-mentioned reacting hole that has encapsulated.Washing then.(doing blank well simultaneously, negative control hole and positive control hole).
3. add the Avidin labelled antibody: in each reacting hole, add antibody (available from U.S. abcam company) (dilutability after the titration) 0.1ml of the Avidin mark of fresh dilution.Hatched 0.5～1 hour washing for 37 ℃.4. add biotin labeled horseradish peroxidase: in each reacting hole, add biotin labeled horseradish peroxidase (available from the U.S. sigma company) 0.1ml of fresh dilution.Hatched 0.5 hour washing for 37 ℃.
5. add substrate solution colour developing: in each reacting hole, add tmb substrate (available from U.S. sigma company) the solution 0.1ml of interim preparation, 37 ℃ 10～30 minutes.
6. cessation reaction: in each reacting hole, add 2M hydrochloric acid 0.05ml.
7. the result judges: can be on white background, and the result directly detects by an unaided eye: color is dark more in the reacting hole, and positive degree is strong more, and negative reaction is colourless or extremely shallow, according to the depth of be color, with "+", "-" number expression.Also can survey the OD value: on the ELISA detector, locate, survey each hole OD value with zeroing back, blank hole in 450nm (if with ABTS colour developing, then 410nm), if it is greater than 2.1 times of the negative control OD value of stipulating, promptly positive.
8. come the production standard curve through the galactose agglutinin 3 (available from U.S. sigma company) that in reacting hole, adds gradient dilution,, calculate the amount of the galactose agglutinin 3 in each sample according to typical curve.Testing result is as shown in the table:
| Time | Control group galactose agglutinin 3 concentration (pg/ml) | Model group galactose agglutinin 3 concentration (pg/ml) |
| The 0th week | 0.26 ± 0.03 | 0.23 ± 0.04 |
| The 1st week | 0.23 ± 0.05 | 0.58 ± 0.07 |
| The 2nd week | 0.28 ± 0.08 | 3.67 ± 0.33 |
| The 3rd week | 0.19 ± 0.02 | 16.71 ± 2.03 |
| The 4th week | 0.24 ± 0.01 | 37.91 ± 6.03 |
Interpretation of result: each time point of experimental group and control group, comprise 10 mouse, through the t check, the p value between two groups is 0.0021, has significant difference.Can detect the galactose agglutinin 3 in the serum easily through enzyme linked immunoassay, the content of the galactose agglutinin 3 in the animal blood of generation liver fibrosis is apparently higher than intact animal.
Claims (2)
1. blood mark that is used for diagnosing liver fibrosis and early-phase hepatocirrhosis, said blood mark is a galactose agglutinin 3.
2. application rights requires 1 described blood mark to carry out the analysis and evaluation of liver fibrosis and early-phase hepatocirrhosis through the concentration of the galactose agglutinin in the enzyme linked immunoassay analyzing blood 3.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2012100947233A CN102645540A (en) | 2012-04-01 | 2012-04-01 | Blood marker for diagnosing hepatic fibrosis and early cirrhosis |
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| Application Number | Priority Date | Filing Date | Title |
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| CN2012100947233A CN102645540A (en) | 2012-04-01 | 2012-04-01 | Blood marker for diagnosing hepatic fibrosis and early cirrhosis |
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| CN102645540A true CN102645540A (en) | 2012-08-22 |
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103513038A (en) * | 2013-05-16 | 2014-01-15 | 武汉生之源生物科技有限公司 | Detection kit for galectin-3 and preparation method thereof |
| CN103808944A (en) * | 2014-03-07 | 2014-05-21 | 高平 | Biomarkers von willebrand factor (VWF) and ADAMTS13 and use thereof in cirrhosis diagnosis reagent |
| CN107942055A (en) * | 2017-11-22 | 2018-04-20 | 南宁科城汇信息科技有限公司 | The ELISA method of transcript profile and protein science in a kind of liver cancer biological process |
| CN108931645A (en) * | 2018-07-26 | 2018-12-04 | 北京大学第医院 | A kind of HCV removes heptic fibrosis assessment system and appraisal procedure |
| CN115902204A (en) * | 2022-09-23 | 2023-04-04 | 北京大学第三医院（北京大学第三临床医学院） | Detection method and kit for exosomal galectin 9 |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1759320A (en) * | 2003-01-14 | 2006-04-12 | Vib研究所 | A serum marker for measuring liver fibrosis |
| CN101356439A (en) * | 2005-07-01 | 2009-01-28 | 西马生物医学计划公司 | Fibrosis markers |
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2012
- 2012-04-01 CN CN2012100947233A patent/CN102645540A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1759320A (en) * | 2003-01-14 | 2006-04-12 | Vib研究所 | A serum marker for measuring liver fibrosis |
| CN101356439A (en) * | 2005-07-01 | 2009-01-28 | 西马生物医学计划公司 | Fibrosis markers |
Non-Patent Citations (6)
| Title |
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| JOSEF WANNINGER,ET AL.: "Systemic and hepatic vein galectin-3 are increased in patients with alcoholic liver cirrhosis and negatively correlate with liver function", 《CYTOKINE》 * |
| NEIL C. HENDERSON，ET AL.: "Galectin-3 regulates myofibroblast activation and hepatic fibrosis", 《PNAS》 * |
| S.HONSAWEK,ET AL.: "Elevation of Serum Galectin-3 and Liver Stiffness Measured by Transient Elastography in Biliary Atresia", 《EUROPEAN JOURNAL OF PEDIATRIC SURGERY》 * |
| 封益飞 等: "半乳糖凝集素-3对肝星状细胞增殖及凋亡的影响", 《中华肝脏病杂志》 * |
| 张鸿 等: "Galectin-3在CCl4所致肝损伤的保护作用", 《解剖科学进展》 * |
| 方青青 等: "肝癌组织和血清中半乳糖血凝素-3的表达及其意义", 《中华肝脏病杂志》 * |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103513038A (en) * | 2013-05-16 | 2014-01-15 | 武汉生之源生物科技有限公司 | Detection kit for galectin-3 and preparation method thereof |
| CN103513038B (en) * | 2013-05-16 | 2015-06-10 | 武汉生之源生物科技有限公司 | Detection kit for galectin-3 and preparation method thereof |
| CN103808944A (en) * | 2014-03-07 | 2014-05-21 | 高平 | Biomarkers von willebrand factor (VWF) and ADAMTS13 and use thereof in cirrhosis diagnosis reagent |
| CN103808944B (en) * | 2014-03-07 | 2016-04-20 | 高平 | Biomarker VWF and ADAMTS13 and the purposes in liver cirrhosis diagnosis reagent thereof |
| CN107942055A (en) * | 2017-11-22 | 2018-04-20 | 南宁科城汇信息科技有限公司 | The ELISA method of transcript profile and protein science in a kind of liver cancer biological process |
| CN108931645A (en) * | 2018-07-26 | 2018-12-04 | 北京大学第医院 | A kind of HCV removes heptic fibrosis assessment system and appraisal procedure |
| CN115902204A (en) * | 2022-09-23 | 2023-04-04 | 北京大学第三医院（北京大学第三临床医学院） | Detection method and kit for exosomal galectin 9 |
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Application publication date: 20120822 |