In-situ transplantation process of constructing human liver cancer mouse model

A technology of mouse model and human liver cancer cell line, which is applied in the field of xenogeneic orthotopic transplantation to construct animal models of human liver cancer, can solve the problem of limiting tumor biology and the research and evaluation of the therapeutic effect of anti-tumor immune drugs, and cannot truly simulate liver cancer tumors Cellular immune response status, feeding and experimental requirements are high, to achieve the effect of prolonging the survival time with tumors, cheap animals, and large capacity to withstand cancerous ascites

Inactive Publication Date: 2007-08-22
SHANDONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0010] Such models have a high success rate of xenotransplantation, short incubation period, but most of them have low metastasis rate
At the same time, due to the lack of an immune system mainly composed of T cells, nude mice require high feeding and experimental conditions, short lifespan, and low tolerance to experimental treatment factors, especially transplanted cells such as liver cancer, which grow rapidly in animals and have a short lifespan. A large number of metastatic nodules and fatal bloody ascites will be produced within a short period of time, which will make the survival time of nude mice with tumors short and affect the experimental research. It is not suitable for dynamic observation in drug-targeted therapy, thus limitin

Method used

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  • In-situ transplantation process of constructing human liver cancer mouse model
  • In-situ transplantation process of constructing human liver cancer mouse model
  • In-situ transplantation process of constructing human liver cancer mouse model

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] Example 1: Preparation of DiI fluorescent dye-labeled human liver cancer cell HepG-2 suspension

[0060] 1. Test equipment

[0061] Fluorescence microscope (NIKON ECLIPSE TE2000-U); cell culture incubator (SANYO, 2300TC); 0.22 μm syringe filter (MILLIPORE, USA);

[0062] 2. Test reagent

[0063] DMEM-H liquid basic culture medium (HyClone); premium fetal bovine serum (HyClone); fluorescent dye DiI (AnaSpec, Inc.); 0.25% trypsin (SINGAMA), etc.

[0064] 3. Test method

[0065] AFP-positive human liver cancer cell line HepG-2, purchased from Shanghai Institute of Cell Biology, Chinese Academy of Sciences, stored in liquid nitrogen, DMEM-H basic culture medium, 10% fetal bovine serum, subcultured in a 37°C 5% CO2 constant temperature incubator; 0.25 % Sterile trypsinization for passage.

[0066] Dissolve the fluorescent dye DiI in absolute ethanol to 2.0 mg / ml; filter and sterilize with a 0.22 μm disposable filter, and store in an airtight and dark place at room temper...

Embodiment 2

[0067] Example 2: Construction of orthotopic transplantation human liver cancer cell HepG-2 mouse model

[0068] 1. Test material

[0069] 30-gauge 50-μl glass microsampler (Shanghai Gaoxin Glass Instrument Factory); DiI fluorescent dye-labeled human liver cancer cell HepG-2 suspension, cryostat (MICROM HM 550); AO 820 microtome (American Optical Corporation ); paraffin (sangon), etc.;

[0070] 2. Test animals

[0071] Pregnant mice of the Kunming breed were purchased from the Experimental Animal Center of Shandong University. They were adapted to feeding in an SPF clean-level experimental animal room and waited for their birth. Three-day-old suckling mice were taken out as tumor-bearing receptors. Both males and females had an average body weight of 1.9g.

[0072] 3. Test method

[0073] 1) Take 3-day-old Kunming suckling mice, and disinfect the abdominal skin with alcohol.

[0074] 2) Clamp the neck of the mouse with the index finger and middle finger to fix the head of...

Embodiment 3

[0093] Example 3: ELISA detection of serum alpha-fetoprotein concentration changes in tumor-bearing mice

[0094] 1. Test material

[0095] Tumor-bearing mouse serum; Alpha-fetoprotein enzyme immunoassay kit (BioCheck, Inc); Alpha-fetoprotein antibody (LABVISON)

[0096] 2. Test method

[0097] (1) Preparation of cleaning solution for work: 20ml / bottle of concentrated cleaning solution, diluted with distilled water 1:20 before use;

[0098] (2) Adding samples: set 1 well of blank control, 2 wells of negative control and 2 wells of positive control, respectively add 50 μl of negative control and 50 μl of samples to be tested in corresponding wells;

[0099] (3) Enzyme addition: Add 1 drop (50 μl) of enzyme marker to each well except the blank control well, shake gently, and seal the plate well with transparent tape;

[0100] (4) Incubation: 30 minutes in a water bath at 37°C;

[0101] (5) Washing: Remove the liquid in the holes, fill each hole with washing liquid, let it st...

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Abstract

The present invention discloses in-situ transplantation process of constructing human liver cancer mouse model. The process includes using Kunming sucking mouse as the tumor carrying acceptor, and injecting the mouse liver lobes with human liver cancer cell suspension to in-situ transplanted tumor carrying model of human liver cancer cell. The process is simple and high in successful rate. The process makes it possible to monitor the mutual effect between liver cancer cell and mouse immune system under normal mouse body immune environment, simulate the whole process of immunological tolerance and antitumor immunity in human body and provide ideal operation platform for the immunobiological medicine screening and pharmacological research.

Description

technical field [0001] The present invention relates to a preparation method of an animal liver cancer model, in particular to a method for constructing a human liver cancer animal model through xenogeneic orthotopic transplantation under the physiological condition that ordinary mice have normal immune function. Background technique [0002] Primary Hepatocellular Carcinoma (PHC) is one of the malignant tumors with the highest degree of malignancy and mortality in the world; the establishment of an ideal animal model of human liver cancer is the basis for the study of liver cancer, screening of anti-tumor drugs and experimental treatment of liver cancer important basis. In previous studies, a variety of animal liver cancer tumor-bearing models have been invented, but due to the differences between humans and animals and many factors in the formation of human liver cancer, it is necessary to establish a model that fully reflects the biological characteristics of human liver ...

Claims

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Application Information

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IPC IPC(8): A61K49/00C12N5/08
Inventor 白增亮王泽张敬平
Owner SHANDONG UNIV
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