Tumorigenicity Testing-Evaluation of tumor models-Brain Case ()

Evaluation of tumor models

Cancer has become a major threat to human public health, and its death rate is second only to cardiovascular disease. According to the latest data from 2016, 14.1 million new patients were diagnosed worldwide in 2012, and 8.2 million died from malignant tumors. In China, the death rate from cancer remains high, with about 7,700 patients dying from cancer every day. Therefore, the study of tumor has become the top priority of modern biological research, and tumor animal models play a crucial role in tumor development and drug research and development. cell derived xenograft (CDX), which involves the inoculation of tumor cells in vitro to immunodeficient mice, is one of the most commonly used models to study tumor cell proliferation and to screen drugs in vivo. Due to the high homology characteristics of the cells in vitro, the model is easy to establish and has good repeatability. However, during the continuous culture and passage in vitro, the tumor heterogeneity is greatly different from the original tumor tissue.

Test method for tumorigenicity

Nude mouse tumor model is a common and commonly used tumor animal model, which plays an important role in the establishment of subcutaneous tumor transplantation model. The inoculation of nude mouse tumor models generally includes cell inoculation and tumor mass inoculation, and the inoculation samples include surgical biopsy specimens, cancerous pleural fluid specimens and cell lines cultured in vitro.
The establishment of transplanted tumor model in nude mice has the advantages of short establishment period, high tumor formation rate, easy operation and low cost. Tumor transplantation experiments in nude mice are often used to test the tumorigenic ability of cells/the anti-cancer effect of anti-tumor drugs.

Preparation for the experiment:

• 1. 0.25% pancreatic enzyme, serum containing medium, PBS or saline
• 2. 1ml syringe, cell counter, alcohol cotton ball

Experimental procedure:

• 1. Rinse the counting plate and cover glass and let dry
• 2. Conventional digestive cells, centrifuge
• 3. Add an appropriate amount of PBS suspensions to make them into single-cell suspensions
• 4. Apply the cell suspension according to a certain dilution
• 5. Cell count
• 6. Take 5*10^6 cells and centrifuge them again to collect cell precipitation
• 7. The cells were suspended with 0.2mlPBS or normal saline and placed into 0.5ml EP tubes, respectively, and sealed with sealing film
• 8. Select nude mice aged 4 to 6 weeks and disinfect the back skin of nude mice
• 9. The cell suspension was absorbed with a 1ml syringe and injected under the skin of nude mice
• 10. After that, the survival of nude mice and the tumor transplantation were observed every day, and the tumor size was measured
• 11. When the early tumor grew to about 1.5~2cm in diameter, the mice were killed by neck removal, the tumor was removed, weighed, measured the size of the tumor and calculated the volume
• 12. Statistical analysis

Matters needing attention

• 1. Cell suspension should be prepared into single-cell suspension during cell resuspension
• 2. Before counting, the cells should be thoroughly mixed and diluted to the appropriate concentration, and the counting must be accurate

In general, to group nude mice into groups in experiments, random grouping should be adopted, and in principle, the more nude mice in each group, the better, but considering the cost of the experiment, 5-10 mice in each group are generally selected. In the pre-experiment, select 3 cells that can be used for statistical analysis, and after the cell count, it is better not to exceed 0.2ml when the cells are suspended with PBS or normal saline. In terms of the number of inoculated cells, generally 5~10x10⁶ cells are selected.

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Tumorigenicity Testing-Evaluation of tumor models-Brain Case