Application of star colloid cell phosphorylation protein PEA-15

An astrocyte, phosphorylated protein technology, applied in the biological field, can solve problems such as the correlation of hepatocellular carcinoma that has not been seen

Inactive Publication Date: 2007-04-25
SHANGHAI INST OF BIOLOGICAL SCI CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] But so far, there is no report about the correlation between astrocyte phosphorylated protein PEA-15 and hepatocellular carcinoma in the prior art

Method used

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  • Application of star colloid cell phosphorylation protein PEA-15
  • Application of star colloid cell phosphorylation protein PEA-15
  • Application of star colloid cell phosphorylation protein PEA-15

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Example 1 , Preparation of protein samples from hepatocellular carcinoma tissue and paracancerous tissue

[0026] Urea, 3-[(3-cholamidopropyl)-diethylammonium]-1-propanesulfonic acid (CHAPS), phenylmethylsulfonyl fluoride (PMSF), dithiothreitol used in this example (DTT) were purchased from Sigma Company.

[0027] In this example, protein samples of cancer tissue and paracancerous tissue of hepatocellular carcinoma were prepared by nonenzymatic sample preparation (NESP), as follows:

[0028]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 ...

Embodiment 2

[0033] Example 2 , Screening of differentially expressed proteins

[0034] 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; cleavable ICAT reagents were purchased from Applied Biosystems Framingham, MA.

[0035] Ammonium persulfate (AP), TEMED, Tri-n-butylphosphat (TBP), PDQuest software, etc. are Bio-Rad products.

[0036] Avidin affinity column was purchased from Applied Biosystems, Framingham, MA.

[0037] LCQ TM Deca XP system and ProteomeX TM Workstation was purchased from Thermo Finnigan Company.

[0038] The composition of the loading buffer used is: 0.6ml of 1mol / L Tris-HCl (pH6.8), 5ml of 50% glycerol, 2ml of 10% SDS, 0.5ml of mercaptoethanol, 1.9ml of distilled water, and a small amount of hexabromophenol blue.

[0039] First, the gel-enhan...

Embodiment 3

[0047] Example 3 , Western blot verification of differential expression of phosphorylated protein PEA-15 in astrocytes

[0048] In order to confirm the differential expression of the phosphorylated protein PEA-15 in astrocytes, the protein samples of cancer tissues and corresponding paracancerous tissues of 10 patients with hepatocellular carcinoma (prepared by NESP method, the other 10 cases except 429 cases in Table 1 ), using the purchased anti-astrocyte phosphorylated protein PEA-15 antibody for Western blot analysis, the specific process is briefly described as follows:

[0049] Take 20 μg protein samples from each sample and use 12% SDS-PAGE to separate, transfer to PVDF membrane (purchased from Amersham Biosciences), and use rabbit anti-human astrocyte phosphorylated protein PEA-15 monoclonal antibody (purchased from Amersham Biosciences) as the primary antibody. Biosource company, 1:1000 dilution), incubate at room temperature for 2 hours, wash three times with TBST ...

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Abstract

The invention sifts the protein with differential expression which is in cancer organises of the hepatocyte cancer and nearby cancer organises to obtain a high expression protein in the cancer organizes of the hepatocyte cancer. The differential expression of the astrocyte phosphating protein PEA-15 between the cancers organises of the hepatocyte cancer and nearby cancer organizes exists indeed is confirmed by the immune imprint experiment. In consideration of the correlation between the astrocyte cell phosphorylation protein PEA-15 and the hepatocyte cancer, the protein can be used as protein molecule mark to detect the hepatocyte cancer.

Description

technical field [0001] The invention belongs to the field of biotechnology, in particular, the invention relates to an application of astrocyte phosphorylated protein PEA-15 as a protein molecular marker for detecting 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 isolate and identi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/574G01N33/577
Inventor 曾嵘周晓李辰袁新雨
Owner SHANGHAI INST OF BIOLOGICAL SCI CHINESE ACAD OF SCI
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