Method for culturing mini-porcine islet cell in vitro

A technology for islet cells and in vitro culture, which is applied in the direction of cell culture active agents, pancreatic cells, artificial cell constructs, etc., can solve the problems of high cell apoptosis rate, poor stability, and low cell viability, so as to improve vitality and reduce cell viability. Effects of Withering Rate, Significant Market Value, and Social Value

Active Publication Date: 2019-02-19
SOUTH CHINA UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The existing in vitro culture technology of minipig islet cells has the defects of high apoptosis rate, low cell viability and poor stability

Method used

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  • Method for culturing mini-porcine islet cell in vitro

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Embodiment 1

[0033] A method for culturing minipig islet cells in vitro, the steps are as follows:

[0034] 1) Prepare a cell growth solution, the cell growth solution includes the first base solution composed of calf serum, F12 culture medium and DMEM culture medium in a volume ratio of 5:100:25, and add Apoptosis inhibitor Z-VAD-FMK 8 μM (based on the total molar mass of all substances in the cell growth medium), insulin-like growth factor 12 μg / L, vascular endothelial growth factor 5 μg / L, fibroblast growth factor 2 μg / L, TGF -β 6 μg / L, insulin 12 μg / L, penicillin 90 U / mL, streptomycin 90 U / mL, transferrin 3 μg / L, first additive 25 μg / L;

[0035] 2) Prepare a cell culture solution, the cell culture solution includes a second base solution consisting of calf serum, F12 culture medium and DMEM culture medium in a volume ratio of 3:90:30, and add Apoptosis inhibitor Z-VAD-FMK 13 μM, insulin-like growth factor 6 μg / L, vascular endothelial growth factor 2 μg / L, fibroblast growth factor 2 μg...

Embodiment 2

[0050] A method for culturing minipig islet cells in vitro, the steps are as follows:

[0051] 1) Prepare a cell growth solution, the cell growth solution includes the first base solution composed of calf serum, F12 culture medium and DMEM culture medium in a volume ratio of 5:100:25, and add Apoptosis inhibitor Z-VAD-FMK 9 μM, insulin-like growth factor 13 μg / L, vascular endothelial growth factor 6 μg / L, fibroblast growth factor 4 μg / L, TGF-β 8 μg / L, insulin 13 μg / L, penicillin 90U / mL, streptomycin 90U / mL, transferrin 3.5μg / L, first additive 28μg / L;

[0052] 2) Prepare a cell culture solution, the cell culture solution includes a second base solution consisting of calf serum, F12 culture medium and DMEM culture medium in a volume ratio of 3:90:30, and add Apoptosis inhibitor Z-VAD-FMK 14 μM, insulin-like growth factor 8 μg / L, vascular endothelial growth factor 3 μg / L, fibroblast growth factor 4 μg / L, TGF-β 13 μg / L, insulin 9 μg / L, penicillin 90U / mL, streptomycin 90U / mL, tra...

Embodiment 3

[0067] A method for culturing minipig islet cells in vitro, the steps are as follows:

[0068] 1) Prepare a cell growth solution, the cell growth solution includes the first base solution composed of calf serum, F12 culture medium and DMEM culture medium in a volume ratio of 5:100:25, and add Apoptosis inhibitor Z-VAD-FMK 10 μM, insulin-like growth factor 15 μg / L, vascular endothelial growth factor 8 μg / L, fibroblast growth factor 5 μg / L, TGF-β 10 μg / L, insulin 15 μg / L, penicillin 90 U / mL, streptomycin 90U / mL, transferrin 4μg / L, first additive 30μg / L;

[0069] 2) Prepare a cell culture solution, the cell culture solution includes a second base solution consisting of calf serum, F12 culture medium and DMEM culture medium in a volume ratio of 3:90:30, and add Apoptosis inhibitor Z-VAD-FMK 16 μM, insulin-like growth factor 10 μg / L, vascular endothelial growth factor 4 μg / L, fibroblast growth factor 5 μg / L, TGF-β 15 μg / L, insulin 10 μg / L, penicillin 90U / mL, streptomycin 90U / mL,...

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Abstract

The invention discloses a method for culturing a mini-porcine islet cell in vitro. The method comprises the following steps that a cell growth liquid is prepared; a cell culture liquid is prepared; amini-porcine islet cell cryopreservation tube is taken out, melted and placed into a centrifuge tube containing the cell growth liquid to be centrifuged, a liquid supernatant is removed, the cell growth liquid is adopted to suspend the cell at the bottom of the centrifuge tube, then the cell is extracted into a first culture dish containing the cell growth liquid, and the first culture dish is putinto an incubator for cultivation; after cultivation, the culture dish and a culture medium are replaced, a second culture dish containing the cell culture liquid is adopted for cultivation, and thenthe culture dish and the culture medium are replaced every 1-3 days, and the cell culture liquid is used as the culture medium all the time. The method for culturing the mini-porcine islet cell in vitro can reduce the cell wilting rate, increase the islet beta cell viability, and the cultured cell has a sensitive glucose stimulating response function, so that the method can meet the requirementsfor studying human islet diseases and has important market value and social value.

Description

technical field [0001] The invention relates to the technical field of animal cell culture, in particular to a method for culturing minipig islet cells in vitro. Background technique [0002] As the earliest animals raised by humans, pigs have the same omnivorous behavior as humans, but are quite different from dogs (carnivorous) and monkeys (vegetarian). Its physiological function, material metabolism, organ morphology and disease mechanism are very similar to human beings. Pigs and humans have a similar cytochrome oxidase P450 system. The important metabolic enzymatic characteristics determine that the biotransformation of drugs in the body is very similar to that of humans. It is an ideal model animal for drug safety evaluation and screening, and human diseases, especially in anti- Oncology drugs, cardiovascular system drugs, skin-penetrating drugs, non-steroidal antibiotics, and digestive system drugs have shown obvious advantages over dogs and non-human primates. Coup...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071
CPCC12N5/0676C12N2500/25C12N2500/76C12N2501/115C12N2501/105C12N2501/734C12N2501/165C12N2501/15
Inventor 谢水林黄黎珍邹芬
Owner SOUTH CHINA UNIV OF TECH
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