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Tissue preservation solution and application thereof

A technology for preserving liquid and tissue, applied in the field of biomedicine, can solve the problem of inability to preserve cell activity and source tissue for a long time, achieve the effect of maintaining cell development characteristics, high cell vitality, and improving success rate

Active Publication Date: 2021-05-28
ACCURATE INT BIOTECHNOLOGY (GUANGZHOU) CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In summary, the current cell preservation system cannot preserve cell activity and the characteristics of the source tissue for a long time, which has become an important obstacle to the success of primary cell culture.

Method used

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  • Tissue preservation solution and application thereof
  • Tissue preservation solution and application thereof
  • Tissue preservation solution and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0032] This example provides a method for preserving and culturing mouse liver tissue. The tissue preservation solution involved includes Advanced DMEM / F12 medium and added factors; the added factors include components at the following final concentrations: N-acetylcysteine, 5 μM ; Indole-3-lactic acid: 100μM; Y-27632 100μM; GDF11100ng / ml; penicillin-streptomycin mixture, 5×; The preservation and cultivation method specifically includes the following steps:

[0033] (1) After the mice were killed, soak them in normal saline for 10 minutes, soak them in alcohol for 30 seconds, take the liver after dissection, and cut it into about 2mm with surgical scissors 3 Place these small pieces in the aforementioned tissue preservation solution and store them at 4°C for 12 hours;

[0034] (2) Then add 10ml collagenase to the tissue preservation solution in step (1) to resuspend, transfer to 37°C, 220rpm shaker for digestion for 20min, filter the cells with a 100um cell mesh, add 10ml DME...

Embodiment 2

[0036] This embodiment provides a method for preserving and culturing mouse intestinal cancer tissue, the tissue preservation solution involved includes Advanced DMEM / F12 medium and added factors; the added factors include the following components at final concentrations: N-acetylcysteine, 2μM; indole-3-lactic acid: 500μM; Y-27632 50μM; penicillin streptomycin mixed solution, 3×; Primocin, 2mg / ml; EGF, 50ng / ml; CHIR99021, 5μM; Mix it evenly into the base medium. The preservation and cultivation method specifically includes the following steps:

[0037] (1) One PDX mouse model of intestinal cancer was taken, sacrificed and dissected to obtain tumor tissue. Surgical scissors shredded to 2mm 3 Place these small pieces in the aforementioned tissue preservation solution and store them at 4°C for 48 hours;

[0038] (2) Isolate tumor cells by slide method, collect tumor cells and resuspend in DMEM / F12 medium, count the cells, and calculate the cell yield, then inoculate the cells ...

Embodiment 3

[0040]This embodiment provides a method for preserving and culturing mouse lung tissue. The tissue preservation solution involved includes Advanced DMEM / F12 medium and added factors; the added factors include components at the following final concentrations: N-acetylcysteine, 0.2 μM; indole-3-lactic acid: 1000μM; Y-27632 10μM; GDF1110ng / ml; penicillin-streptomycin mixture, 1×; . Preservation and cultivation method specifically comprises the following steps:

[0041] (1) After the mice were killed, they were soaked in normal saline for 10 minutes, soaked in alcohol for 30 seconds, and the lungs were taken after dissection, and the bronchi were stripped as much as possible, and the lung parenchyma was harvested, and cut into about 1mm with surgical scissors 3 Place these small pieces in the aforementioned tissue preservation solution and store them at 4°C for 12 hours;

[0042] (2) Then add 10ml collagenase to the tissue preservation solution in step (1) to resuspend, transfer...

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Abstract

The invention provides a tissue preservation solution and an application thereof. The tissue preservation solution comprises a basic culture solution and additive factors, the addition factor at least comprises the following components with final concentration: N-acetylcysteine with the concentration of 0.2 to 5 [mu]M; indole-3-lactic acid with the concentration of 100 to 1000 [mu]M; Y-27632 with the concentration of 10 to 100 [mu]M; GDF11 with the concentration of 10 to 100 ng / ml; a penicillin-streptomycin mixed solution with 1-5 *; and Primocin with the concentration of 0.5 to 5 mg / ml. The tissue preservation solution is applied to preservation of in-vitro tissues before primary culture. The preservation method comprises the following steps: completely immersing tissues in vitro into the tissue preservation solution, and preserving at 2-8 DEG C. According to the tissue preservation solution, the effective preservation period of normal tissues after in-vitro treatment exceeds 12 hours, the effective preservation period of tumor tissues and cells after in-vitro treatment exceeds 48 hours, the tissues preserved for a long time can still keep high cell viability and keep cell development characteristics, and the success rate of primary cell culture is remarkably increased. In addition, the tissue preservation solution disclosed by the invention has important significance on primary culture specimens needing to be preserved for a long time, especially precious clinical specimens.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to a tissue preservation solution and its application. Background technique [0002] Primary culture is an important research method, including adherent culture, suspension culture, 3D culture and other methods. However, the success rate of primary culture has a great correlation with the activity of source tissue samples. However, due to the influence of various factors after the tissue is isolated, the cell viability decreases or even dies, which makes it impossible to obtain a sufficient amount of active cells during the pretreatment process of the primary culture, which leads to the failure of the culture. For example, the loss of contact with other cells in vivo or extracellular mechanisms induces the regulation of related apoptotic mechanisms after cells are isolated, resulting in anoikis. In addition, after organoids are isolated from the body, due to the lo...

Claims

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Application Information

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IPC IPC(8): A01N1/02
CPCA01N1/021A01N1/0226
Inventor 陈泽新朱宇李刚黄敏
Owner ACCURATE INT BIOTECHNOLOGY (GUANGZHOU) CO LTD
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