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Three-dimensional cell cultivator based on temperature-responsive liquid marbles and construction and application methods of three-dimensional cell cultivator

A temperature-responsive, three-dimensional cell technology, used in tissue cell/virus culture devices, biochemical equipment and methods, and methods for supporting/immobilizing microorganisms. The overall microenvironment required for growth and differentiation, the lack of three-dimensional scaffolds, etc., achieve the effects of excellent stability and gas permeability, easy mass production, and broad application prospects.

Active Publication Date: 2017-12-29
NORTHWESTERN POLYTECHNICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Up to now, a large number of techniques for obtaining cell sheets by PNIPAm and its copolymers in two-dimensional surface modified cell culture media have been developed, but the CST cell sheet technology still stays at the traditional two-dimensional cell culture level. The three-dimensional scaffold composed of the matrix cannot form the overall microenvironment required for cell growth and differentiation, and lacks the specific growth factors and differentiation factors in the microenvironment in the body, so the cells attached to the substrate plane can only expand two-dimensionally. How to realize the 3D cell culture of cell layer technology is a great challenge

Method used

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  • Three-dimensional cell cultivator based on temperature-responsive liquid marbles and construction and application methods of three-dimensional cell cultivator
  • Three-dimensional cell cultivator based on temperature-responsive liquid marbles and construction and application methods of three-dimensional cell cultivator
  • Three-dimensional cell cultivator based on temperature-responsive liquid marbles and construction and application methods of three-dimensional cell cultivator

Examples

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Effect test

example 1

[0032] (1) Preparation of hybrid particles

[0033] When the inorganic micro-nanoparticles are nano-silica, and the temperature-sensitive polymer is poly-N-isopropylacrylamide PNIPAM homopolymer, use 3-aminopropyltriethoxysilane and 2-bromoisobutyryl bromide with SiO 2 reaction to get SiO 2 -Br initiator, and then use surface-initiated atom transfer radical polymerization to connect PNIPAm to SiO 2 surface, to obtain SiO with a surface grafting rate of 40% 2 @PNIPAm.

[0034] (2) Construction of liquid marble three-dimensional cell culture device

[0035]With Hep G2 (approximately 10 per 100 μL 4 ) The cell culture medium is (DMEM medium (Gibco), 10% calf serum, and penicillin solution 100U, NaHCO 3 buffer solution) was used as a cell culture medium and temporarily stored at 5°C. Spread the synthesized temperature-sensitive hybrid particles on the plastic substrate, add 150 μL of cell culture solution dropwise with a micro-injector, and then use the micro-injector to dr...

example 2

[0039] (1) Preparation of hybrid particles

[0040] When the inorganic micro-nanoparticles are nano-silica, and the temperature-sensitive polymer is PNIPAm / PAA, a copolymer of poly-N-isopropylacrylamide and polyacrylic acid, 3-aminopropyltriethoxysilane and 2- Bromoisobutyryl bromide and SiO 2 reaction to get SiO 2 -Br initiator, and then graft PNIPAm and PAA monomers to SiO by initiating atom transfer radical polymerization on the graft surface 2 surface, to obtain a surface grafting rate of 40% of SiO 2 @PNIPAM / PAA particles.

[0041] (2) Construction of liquid marble three-dimensional cell culture device

[0042] With Hep G2 (approximately 10 per 100 μL 4 ) The cell culture medium is (DMEM medium (Gibco), 10% calf serum, and penicillin solution 100U, NaHCO 3 buffer solution) was used as a cell culture medium and temporarily stored at 5°C. Spread the synthesized temperature-sensitive hybrid particles on the plastic substrate, add 150 μL of cell culture solution dropwi...

example 3

[0046] (1) Preparation of hybrid particles

[0047] When the temperature-sensitive polymer is nano-ferric oxide, the silane coupling agent 2-(4-chlorosulfonylphenyl) ethyl trichlorosilane (CTCS) is grafted onto Fe 3 o 4 surface, and then the PNIPAM / PEG copolymer was grafted onto Fe by surface-initiated atom transfer radical polymerization 3 o 4 surface, to obtain a surface grafting rate of 30% Fe 3 o 4 @PNIPAm / PAA Particles.

[0048] (2) Construction of liquid marble three-dimensional cell culture device

[0049] With Hep G2 (approximately 10 per 100 μL 4 ) The cell culture medium is (DMEM medium (Gibco), 10% calf serum, and penicillin solution 100U, NaHCO 3 buffer solution) was used as a cell culture medium and temporarily stored at 5°C. Spread the synthesized temperature-sensitive hybrid particles on the plastic substrate, add 120 μL of cell culture solution dropwise with a micro-injector, and then use the micro-injector to drive the droplet to roll, so that the surf...

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Abstract

The invention relates to a three-dimensional cell cultivator based on temperature-responsive liquid marbles and construction and application methods of the three-dimensional cell cultivator. For the three-dimensional cell cultivator incorporating the liquid marbles, different types of particles modified by temperature-sensitive copolymers are used as the temperature-response particles, on which different cell culture fluid is dropwise added to form liquid marbles having response behaviors to different temperatures, so that the liquid marbles can serve as the three-dimensional cell cultivator. The three-dimensional cell cultivator can have broad application prospect in the field of biomedical cell culture. The three-dimensional cell cultivator disclosed by the invention has the advantages that a preparation method is quick and mass production is easy to realize; a spherical cultivator of the liquid marbles provides a good three-dimensional template for adhering and proliferating of cells; a temperature controlling method is adopted to realize the cultivation of a complicated cell layer structure and a non-destructive desorption process; the liquid marbles have excellent stability and gas permeability, and can effectively ensure the survival of the cells.

Description

technical field [0001] The invention belongs to the field of colloid and interface science, and relates to a three-dimensional cell culture device based on temperature-responsive liquid marbles and its construction and use method. The spherical culture device of the marbles provides a good three-dimensional template for cell adhesion and proliferation. The method can realize the cultivation of complex cell layer structure and the process of non-destructive desorption, and the liquid marble has excellent stability and gas permeability, which can effectively ensure the survival of cells. Background technique [0002] Traditional cell recovery methods mainly include enzymatic and mechanical methods, but both methods damage membrane proteins to some extent and cause cell function loss. Therefore, the development of non-enzymatic and non-mechanical methods to separate cells from the culture mechanism is current research hotspot. In recent years, the use of temperature-sensitive ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C08F292/00C08F220/54C08F220/06C12M3/00C12M1/00
CPCC08F292/00C12M25/04C12N2513/00C08F220/54C08F220/06
Inventor 陈芳陈闯陈助国马晓燕
Owner NORTHWESTERN POLYTECHNICAL UNIV
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