Method for in-situ cryopreservation of cells by three-dimensional micro-carrier

A cryopreservation and microcarrier technology, applied in the field of cell culture, can solve the problems affecting the subsequent culture of cells, inability to realize in situ cryopreservation, interruption of three-dimensional culture, etc., to achieve long-term preservation, convenient storage, and unchanged cell characteristics Effect

Active Publication Date: 2019-04-05
BEIJING CYTONICHE BIOTECH CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In the prior art, after cell culture, the cells need to be separated from the microcarriers and then frozen for preservation, which will interrupt the process of three-dimensional culture and affect the subsequent culture of cells
Commonly used competitive microcarriers will rupture after cryopreservation, and after thawing, cells fall off from the microcarriers, making in situ cryopreservation impossible

Method used

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  • Method for in-situ cryopreservation of cells by three-dimensional micro-carrier
  • Method for in-situ cryopreservation of cells by three-dimensional micro-carrier
  • Method for in-situ cryopreservation of cells by three-dimensional micro-carrier

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Effect test

Embodiment 1

[0037] 1. Centrifuge the three-dimensional microcarrier suspension containing HEK293T cells at 400×g for 2 minutes, discard the supernatant.

[0038] 2. Add cryopreservation solution (90% FBS+10% DMSO) and divide into cryopreservation tubes (10 mg of cell-containing carrier in 1 ml of freezing solution, 0.5-1 ml per tube).

[0039] 3. Put the cryopreservation tube into the cell program cooling box, put it in the -80 ℃ refrigerator within 5 minutes, and transfer it to liquid nitrogen after 24 hours.

[0040] 4. Recovery of cells adsorbed on microcarriers: When recovering, first prepare a 37°C pre-warmed basal medium, and then quickly freeze the tube in a 37°C water bath, shaking it constantly during the period, until the remaining 1 grain of rice-sized ice cube When (about 1-2 minutes water bath), quickly remove from the water bath. Then dilute the thawed cell-containing three-dimensional microcarrier suspension with basal medium at a dilution ratio of 1:5, then centrifuge at ...

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Abstract

The invention discloses a method for in-situ cryopreservation of cells by a three-dimensional micro-carrier. The method includes the following steps: (1) centrifuging a three-dimensional micro-carriersuspension of cells, and discarding a supernatant to obtain a carrier containing cells; (2) mixing the carrier containing the cells with a cryopreservation solution, and then adding the mixture intoa cryopreservation tube; (3) cooling the cryopreservation tube added with the carrier containing the cells in the step (2), and then transferring to liquid nitrogen to realize in-situ cryopreservationof the cells by the three-dimensional micro-carrier. The method can realize in-situ cryopreservation of cell culture on the three-dimensional micro-carrier without separating the cells from the micro-carrier and preforming cryopreservation, realizes that cell cryopreservation does not interrupt the three-dimensional culture process, is convenient for users to store, transport and resuscitate thecultured three-dimensional cell products, and is further convenient for long-term preservation of the cells.

Description

technical field [0001] The invention relates to a method for in-situ cryopreservation of cells by three-dimensional microcarriers, belonging to the technical field of cell culture. Background technique [0002] With the rapid development of interdisciplinary subjects such as biomedicine, materials science, mechanics, and engineering, more and more people are paying attention to the difference between the microenvironment of cells cultured in vitro and the microenvironment in vivo. The traditional two-dimensional cell culture (based on commercial culture dishes or multi-well plates) technology has been developed for more than 100 years, and it has been widely used in basic life science research, medical research and other fields. However, with the rapid development of microscopic imaging technology, researchers have gradually discovered the living state of cells in a two-dimensional culture environment, and the cell morphology is far from the culture conditions in the real en...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01N1/02
CPCA01N1/0221
Inventor 鄢晓君刘伟
Owner BEIJING CYTONICHE BIOTECH CO LTD
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