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High-flux cultural method for organoid type spheroid

A culture method and high-throughput technology, applied in the field of high-throughput culture of organoid spheroids, can solve the problems of uneven and uncontrollable organoid preparation, inaccurate cell line culture results, inconvenient organoid distribution, etc., to achieve Drug susceptibility testing and toxicity testing, high application value, and the effect of accurate results

Active Publication Date: 2019-07-23
TSINGHUA BERKELEY SHENZHEN INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] In view of the deficiencies of the prior art, the purpose of the present invention is to provide a high-throughput culture method of organoids to solve the problems of inaccurate cell line culture results, uneven and uncontrollable preparation of organoids, and inconvenient distribution of organoids

Method used

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  • High-flux cultural method for organoid type spheroid
  • High-flux cultural method for organoid type spheroid
  • High-flux cultural method for organoid type spheroid

Examples

Experimental program
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Effect test

Embodiment 1

[0056] This embodiment provides a high-throughput culture method for organoid spheroids, comprising the following steps:

[0057] (1) Use the primary cell extraction method to extract primary cells, and count the cells; 2.0×10 6 Mix one primary cell with 100 μL matrigel, inject it into a 1mL syringe, inject fluorine oil into another syringe, place the fluorine oil and matrigel in two syringe pumps and put them in a 4°C refrigerator; The 800μm polytetrafluoroethylene (PTFE) tube, the two tubes merge into the polydimethylsiloxane (PMDS) three-way device, and then the three-way device is connected to another PTFE tube with a diameter of 800μm, and the entire platform is placed at 4 ℃ refrigerator; the flow rate of the injection pump containing Matrigel is 20 μL / min, and the flow rate of the fluorine-containing oil injection pump is 50 μL / min to generate uniform organoid spheroids;

[0058] (2) Connect the organoid spheroids to the 3D printing platform through a 37°C heating devi...

Embodiment 2

[0061] This embodiment provides a high-throughput culture method for organoid spheroids, comprising the following steps:

[0062] (1) Use the primary cell extraction method to extract primary cells, and count the cells; 5.0×10 6 Mix one primary cell with 200μL Matrigel, inject it into a 1mL syringe, inject fluorine oil into the other syringe, place the fluorine oil and Matrigel in two syringe pumps and put them in a 4°C refrigerator; The 800μm polytetrafluoroethylene (PTFE) tube, the two tubes merge into the polydimethylsiloxane (PMDS) three-way device, and then the three-way device is connected to another PTFE tube with a diameter of 800μm, and the entire platform is placed at 4 ℃ refrigerator; the flow rate of the injection pump containing Matrigel is 25 μL / min, and the flow rate of the fluorine-containing oil injection pump is 55 μL / min to generate uniform organoid spheroids;

[0063] (2) Connect the organoid spheroids to the 3D printing platform through a 37°C heating dev...

Embodiment 3

[0066] This embodiment provides a high-throughput culture method for organoid spheroids, comprising the following steps:

[0067] (1) Use the primary cell extraction method to extract primary cells, and count the cells; 1.0×10 6 1 primary cell and 80 μL of Matrigel were mixed thoroughly, injected into a 1mL syringe, and the other syringe was injected with fluorine oil. The fluorine oil and Matrigel were respectively placed in two syringe pumps and placed in a 4°C refrigerator; the syringes were respectively connected with diameters It is a 600 μm polytetrafluoroethylene (PTFE) tube, and the two tubes are merged into a polydimethylsiloxane (PMDS) three-way device, and then the three-way device is connected to another PTFE tube with a diameter of 600 μm, and the entire platform is placed at 4 ℃ refrigerator; the flow rate of the injection pump containing Matrigel is 15 μL / min, and the flow rate of the fluorine-containing oil injection pump is 45 μL / min to generate uniform organo...

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Abstract

The invention provides a high-flux cultural method for an organoid type spheroid. The high-flux cultural method comprises the following steps of (1) respectively injecting cell-containing matrigel andfluorocarbon oil into a tee device to obtain the organoid type spheroid; and (2) heating the organoid type spheroid obtained in the step (1), linking the heated organoid type spheroid with a 3D printing platform, distributing the organoid type spheroid, and then culturing the organoid type spheroid. According to the high-flux cultural method provided by the invention, through micro-fluidic, an organoid is obtained, through combination with the 3D printing platform, the organoid can be simply, quickly and accurately printed to a culture carrier, the size and the structure of the organoid are appropriate and uniform and controllable, the number of the organoids in a culture cavity of each culture carrier is unchanged, and besides, drug susceptibility test and toxicity test are realized. Clinical pretest is accurate, the application prospect is favorable, and the high-flux cultural method has high application value.

Description

technical field [0001] The invention belongs to the field of medical devices and relates to a high-throughput culture method for organoid spheroids. Background technique [0002] Cancer has always been the most important factor affecting human health around the world. In recent years, more and more cancer-related targeted new drugs and new tumor immunotherapies have been developed. However, research with patients as research objects is uncontrollable, and the experimental results differ greatly from human to experimental animals, and the migration of experimental results is difficult. 2D cell line culture lacks the in vivo microenvironment and loses some of the characteristics of the tumor, so the results are inaccurate. At present, these problems lead to extremely low conversion efficiency from basic to clinical, and the rise of tumor organoids provides a new technical platform for translational medicine. Tumor organoids are organoids cultured from a single tumor sample. ...

Claims

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Application Information

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IPC IPC(8): C12N5/00
CPCC12N5/0062
Inventor 马少华蒋盛威
Owner TSINGHUA BERKELEY SHENZHEN INST
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