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Protein disulfide bond isomerase A3 and use of antibody thereof in liver cancer detection

A technology of disulfide isomerase and protein is applied in the field of biotechnology detection to achieve the effect of reducing false positive rate, good specificity and improving accuracy

Inactive Publication Date: 2008-12-17
上海中科新生命生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0010] So far, there has been no report of protein disulfide isomerase A3 being associated with hepatocellular carcinoma

Method used

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  • Protein disulfide bond isomerase A3 and use of antibody thereof in liver cancer detection
  • Protein disulfide bond isomerase A3 and use of antibody thereof in liver cancer detection
  • Protein disulfide bond isomerase A3 and use of antibody thereof in liver cancer detection

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1 , Preparation of protein samples from hepatocellular carcinoma tissue and paracancerous tissue

[0027] The urea used in this example, 3-[(3-cholamidopropyl)-diethylammonium]-1-propanesulfonic acid (CHAPS), sodium dodecylsulfonate (SDS), dithiothreose Alcohol (DTT) was purchased from Sigma.

[0028] In this example, protein samples of cancer tissue and paracancerous tissue of hepatocellular carcinoma were prepared by nonenzymatic sample preparation (NESP), as follows:

[0029] The surgically excised fresh tissue block is quickly placed on ice and quickly cut into several small pieces that are visible to the naked eye without areas of necrosis. Use pre-cooled glutamine-free RPMI1640 medium (5% fetal bovine serum, 0.2mM PMSF, 1mM EDTA, benzisoxazole penicillin 25mg / mL, gentamicin 50mg / mL, penicillin 100U / mL, Streptomycin 100mg / mL, Amphotericin B 0.25mg / mL, Nystatin 50U / mL) After washing the small tissue pieces several times, they were quickly ground into cel...

Embodiment 2

[0035] Example 2 , HepG2 and L02 cell line samples incorporating C13-labeled leucine

[0036] The C13-labeled leucine used in this example was purchased from Cambridge Isotope Laboratories, Inc; the dialyzed fetal bovine serum was purchased from hyclone; the D9785 medium was purchased from Sigma.

[0037] The hepatocellular carcinoma cell line HepG2 and the normal liver cell line L02 were cultured for more than six generations in the D9785 medium (20% dialyzed fetal bovine serum) mixed with C13-labeled leucine, transferred to a 10 cm culture dish, and the cells were When it is 80% full, wash it three times with 4°C pre-cooled phosphate buffer on ice, lyse it with an appropriate amount of lysate (8mol / L urea, 4% CHAPS, 40mmol / L Tris and 65mmmol / LDTT), and scrape it with a cell scraper. After scraping, ultrasonic cell breaker (Soniprep 150, UK, MSE) intermittently sonicated for 2 min in an ice bath, and centrifuged at 15000 r / min at 4°C for 1 h. The supernatant was taken, and t...

Embodiment 3

[0038] Example 3 , Using gel electrophoresis, in-gel enzymatic digestion and mass spectrometry to screen differentially expressed proteins in hepatocellular carcinoma tissues and paracancerous tissues

[0039] Urea, 3-[(3-cholamidopropyl)-diethylammonium]-1-propanesulfonic acid (CHAPS), sodium dodecylsulfonate (SDS), dithiothreitol used in this example (DTT) was purchased from Sigma; iodoacetamide (IAA), acrylamide, N, N-methylenebisacrylamide, etc. were purchased from Fluka.

[0040] Ammonium persulfate (IAA), TEMED, PDQuest software, etc. are Bio-Rad products.

[0041] LCQ TM Deca XP system and ProteomeX TM Workstation was purchased from Thermo Finnigan Company.

[0042] Take 5 pairs of the 16 pairs of hepatocellular carcinoma cancer tissue and paracancerous tissue protein samples obtained in Example 1, and take the HepG2 and L02 cell line samples mixed with C13-labeled leucine obtained in Example 2, combine with the coagulation Gel electrophoresis, in-gel enzymatic ...

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Abstract

The invention verifies that protein disulfide isomerase A3 expression in the cancer tissues and adjacent tissues of hepatocellular carcinoma is different through screening the protein having differential expression in the cancer tissues and adjacent tissues of hepatocellular carcinoma and carrying out western blot experiment. The invention discloses the application of the protein disulfide isomerase A3 to the detection of liver cancer and / or susceptibility of liver cancer and the application of the antibody of the protein disulfide isomerase A3 to the detection of liver cancer and / or susceptibility of liver cancer. Therefore, as the potential marker of liver cancer, the protein disulfide isomerase A3 can serve as a marker of the prognosis molecule of liver cancer and a target molecule of clinical treatment.

Description

technical field [0001] The invention belongs to the field of biotechnology detection. Specifically, the invention relates to the application of protein disulfide bond isomerase A3 and its antibody in the detection of liver cancer. Background technique [0002] Liver cancer is a disease that seriously endangers human beings. The incidence of liver cancer in western developed countries is low, and the basic research on liver cancer in the world is still relatively weak. However, my country is a country with a high incidence of liver cancer, and the morbidity and mortality are on the rise, and the age of onset is younger. Medical expenses have increased greatly, and liver cancer has become the number one enemy that seriously endangers the safety of people's life and property in our country, and is an important factor affecting social and economic development. It is of strategic significance to intensify the basic research of liver cancer in my country, and to separate and identi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/533G01N33/563G01N33/574G01N33/577
Inventor 陈薇李辰周晓
Owner 上海中科新生命生物科技有限公司
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