Method and reagent kit for detecting two proteins expressed in human liver organization

A technology of liver tissue and detection methods, applied in biological testing, measuring devices, material inspection products, etc., can solve unclear problems and achieve the effect of wide practicability

Inactive Publication Date: 2008-05-07
INST OF RADIATION MEDICINE ACAD OF MILITARY MEDICAL SCI OF THE PLA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

But how and in what order these factors act at the molecular level is not well understood 2

Method used

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  • Method and reagent kit for detecting two proteins expressed in human liver organization
  • Method and reagent kit for detecting two proteins expressed in human liver organization
  • Method and reagent kit for detecting two proteins expressed in human liver organization

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1 Western blot detection of HOP and nuclear heterogeneous ribosomal protein C1 / C2 in 12 cases of human hepatocellular carcinoma tissues

[0042] 1.1 Put 0.2 g each of the tumor tissues and adjacent non-tumor tissues of 12 patients with hepatocellular carcinoma in liquid nitrogen, and grind them into powder form with a liquid nitrogen precooled mortar. Then the sample was transferred to 1 ml of lysate (7M urea, 2M thiourea, 4% CHAPS, 30mM Tris, pH8.5, protease inhibitor cocktail), and homogenized on ice for 10-20 times. Sonicate in an ice bath for 10s (over 0.1s, stop for 1.9s), and centrifuge at 12,000rpm (20627g) at 4°C for 30min to remove insoluble substances. Protein concentration was determined by Bradford method.

[0043] 1.2 Take 25 μg of each protein sample for SDS-PAGE separation. After the electrophoresis is completed, transfer the protein from the gel to the PVDF membrane with a constant current, current (mA) = membrane area (cm 2 )×2, electric tran...

Embodiment 2

[0044] Example 2 Immunohistochemical detection of HOP and nuclear heterogeneous ribosomal protein C1 / C2 in 70 cases of human hepatocellular carcinoma tissues

[0045] 2.1 The tumor tissues and adjacent non-tumor tissues of 70 patients with hepatocellular carcinoma embedded in paraffin were sliced, and the tumor tissues and adjacent non-tumor tissues of each patient were subjected to immunohistochemical detection in parallel on the same slice.

[0046] 2.2 Paraffin sections were dewaxed and hydrated with 3% H 2 o 2 Incubate at room temperature for 10 minutes to eliminate the activity of endogenous peroxidase. Rinse with distilled water, soak in PBS for 5 min x 3 times, antigen heat recovery, 10% BSA (diluted in PBS) to block, incubate at room temperature for 30 min, pour off the serum, add 1:100 diluted HOP or heterogeneous ribosomal protein C1 / C2 dropwise anti-working solution, and incubated at 37°C for 2h. Rinse with PBS for 5 min x 3 times, add horseradish peroxidase-labe...

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Abstract

The invention relates to a detection method for testing a protein HOP and a heterogeneous ribosome protein C1/C2 which are both expressed in human liver tissue, as well as reagent box used in the method. The method is characterized in that: by means of the fluorescence differential two-dimensional electrophoresis method -MALDI-TOF/TOF, the inventor manages to identify the overexpression of the two proteins on human liver cell carcinoma tumor tissue, and carries out immune histochemistry validations of Western bolt and large amount of samples over the expression levels in the human liver cell carcinoma tissue. The detection method for proteins and the reagent box can be used in the detections for clinical or research purposes.

Description

technical field [0001] The invention relates to an immunohistochemical detection method; specifically, it relates to the detection of two proteins HOP (Hsp70 / Hsp90-organizing protein) and nuclear heterogeneous ribosomal protein C1 / C2 which are up-regulated in human hepatocellular carcinoma tumor tissue (Heterogeneous nuclear ribonucleoprotein C1 / C2, hnRNP C1 / C2) detection method, and a kit for the method. Background technique [0002] Hepatocellular carcinoma is one of the malignant tumors worldwide, with high morbidity and mortality. In 2000, there were 560,000 new cases and almost the same number of deaths 1 . HCC has a higher prevalence in Asia and Africa, and the highest mortality rate in China. Considering that there is still a lack of effective diagnosis and treatment methods for HCC, it is still urgent and necessary to discover important cancer-related molecules of HCC. [0003] Multiple risk factors for the development of HCC, such as HBV / HCV infection, alcohol, a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68G01N33/574
Inventor 贺福初孙薇邢宝才
Owner INST OF RADIATION MEDICINE ACAD OF MILITARY MEDICAL SCI OF THE PLA
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