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A method for cryopreserving rat pancreatic islet cells using microchannels

A technology of islet cells and microchannels, which can be used in applications, preservation of human or animal bodies, animal husbandry, etc. to achieve the effects of reducing dosage, reducing osmotic damage, and reducing damage

Inactive Publication Date: 2018-01-23
WUHAN UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The present invention is based on low-molecular-weight amino acid gelatin factor, which uses its self-assembled supramolecular gel to embed rat islet cells in the confined space of microchannels, and then cryopreserves them, but there is no literature on this aspect to report

Method used

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  • A method for cryopreserving rat pancreatic islet cells using microchannels
  • A method for cryopreserving rat pancreatic islet cells using microchannels
  • A method for cryopreserving rat pancreatic islet cells using microchannels

Examples

Experimental program
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Effect test

Embodiment 1

[0031] Embodiment 1: the preparation of microchannel device

[0032] Such as figure 1 and figure 2 As shown, the microchannel device is composed of a microchannel 5 and a slide glass 6, the microchannel 5 is glued on the slide glass 6, and the length, width and height of the microchannel device are 75mm, 25mm and 5mm respectively. The microchannel 5 includes four parts: a microchannel inlet 1 , a microchannel inlet 2 , a microchannel pipe 3 , and a microchannel outlet 4 . Microchannel pipeline 3 is the main part of microchannel 5, is positioned at the lower surface of microchannel 5, contacts with the upper surface of glass slide 6, and microchannel pipeline 3 communicates with microchannel inlet 1, microchannel inlet 2 and microchannel outlet 4 , the microchannel pipe 3 has a width of 280 μm and a depth of 220 μm.

[0033] The microchannel inlet 1 is located on the left side of the microchannel device, and communicates with the microchannel pipeline 3, and is used to intr...

Embodiment 2

[0036] Embodiment 2: preparation and performance of supramolecular gel

[0037] 1. According to the method described in the literature [10] (Ma Xiaodan. Synthesis of tyrosine derivatives and their gel properties [D]. Xi'an University of Science and Technology, 2012.), the gel factor G was synthesized. The gel factor G can self-assemble in the confined space of the microchannel to form a tight fibrous three-dimensional network structure. On the one hand, this three-dimensional network structure can limit the growth of ice crystals and reduce the mechanical damage caused by the formation of ice crystals to cells; On the other hand, the tight three-dimensional network structure of the supramolecular gel can play a slowing effect, greatly reducing the permeation rate of the cryoprotectant, and reducing the damage caused by the sudden change of the osmotic pressure difference between the inside and outside of the cell membrane.

[0038] 2. Add gel factor G to RPMI1640 medium, and d...

Embodiment 3

[0041] Embodiment 3: the cryopreservation process of rat islet cells

[0042] 1. Acquisition of rat islet cells: Collect rat islet cells from rats purchased by the Hubei Provincial Center for Disease Control and Prevention and put them into RPMI1640 medium containing fetal bovine serum (the volume ratio of fetal bovine serum to RPMI1640 medium is 1: 9), and then placed at 37°C, CO 2 Cultured in an incubator with a volume fraction of 5%.

[0043] 2. Preparation of rat islet cell suspension S2 containing gel factor G:

[0044] ①Take the above-mentioned cultured rat islet cells, digest them with 0.1% trypsin solution and blow them into a single cell suspension, and suspend the cells in RPMI1640 medium containing fetal bovine serum (fetal bovine serum and RPMI1640 medium The volume ratio is 1:9) for later use.

[0045]② Add gel factor G to RPMI1640 medium, and dissolve it completely by ultrasonication at 50°C for 6 minutes to prepare a solution containing gel factor G, and then...

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Abstract

The invention provides a method for utilizing a micro-channel for cryopreserving rat islet cells at low temperature. Restricted space of the micro-channel is utilized for making a three-dimensional structure of supermolecule gel changed, so that the rat islet cells are well protected. Specifically, gelator with the low molecular weight is dissolved into a cell culture medium, after the gelator is dissolved, rat islet cell suspension liquid containing the gelator is prepared; the suspension liquid is guided into the micro-channel and placed into an ice-water bath at the temperature of 3 DEG C-5 DEG C to be gelatinized, then a cryopreserving protective agent is guided into the micro-channel, and slowly permeates into the cells, and finally, the micro-channel device is arranged in a programmed freezing box to be placed into a refrigerator at the temperature of minus 75 DEG C-minus 85 DEG C to be cryopreserved. The new method is provided for freezing the rat islet cells. The freezing point of a freezing system is lowered, freezing damage to the rat islet cells in the freezing process is reduced, the survival rate of the rat islet cells is increased, the activity and the function of the rewarmed rat islet cells are maintained, and the method has a wide application prospect.

Description

technical field [0001] The invention relates to the technical field of cryopreservation of cells, in particular to a method for cryopreserving rat islet cells using a microchannel. It self-assembles in the confined space of the microchannel to form a supramolecular gel with a fibrous three-dimensional network structure, thereby protecting rat islet cells during cryopreservation and reducing cell cryopreservation damage. Background technique [0002] Diabetes is a common endocrine disease. According to data released by the International Diabetes Federation, there are currently 382 million people with diabetes worldwide, and the number of people with diabetes in my country ranks first in the world, accounting for one-third of the total number of patients in the world. Cell transplantation is an effective means of treating diabetes. However, since the number of living cells obtained each time is difficult to meet the needs of a clinical transplantation, the preservation of isl...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A01N1/02
CPCA01N1/0221A01N1/0242
Inventor 陈万煜张丽李鹏程陈熹
Owner WUHAN UNIV OF TECH
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