Breast cancer organoid culture kit

An organoid, breast cancer technology, applied in cell culture active agent, culture process, tissue culture, etc., can solve the problems of error, contamination, preparation, complicated operation steps, etc.

Pending Publication Date: 2020-08-07
扈晖
View PDF0 Cites 12 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The general organoid culture method is based on the construction of cell line cells, or in vitro culture using tumor tissue from animal models. The whole process is relatively complicated, and more than a dozen cytokines and regulatory factors need to be added to the culture medium used. Both need to be dissolved and prepared during use. On the one hand, pollution and preparati...

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Breast cancer organoid culture kit
  • Breast cancer organoid culture kit
  • Breast cancer organoid culture kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] 1. Preparation of the kit

[0030] Preparation of DMEM / F12 medium: DMEM medium is a medium containing amino acids and glucose with high nutrient concentration, while F12 medium has complex components and contains a variety of trace elements. For DMEM / F12 medium. Suitable for the organoid culture of the present invention.

[0031] Preparation of sample preservation solution: take 1000ml of DMEM / F12 medium combined at 1:1, add 0.5ml of prepared penicillin solution (take 800,000 U / bottle of penicillin, dissolve in 4ml of sterile distilled water) to make the final concentration is 100U / ml; add 0.5ml of prepared streptomycin solution (take 1g / bottle of streptomycin and dissolve it in 5ml sterile distilled water) to make the final concentration 0.1mg / ml; add prepared L-Valley 10ml of aminoamide solution (2.922g of L-glutamine was dissolved in three distilled water and added to 100ml to make a solution of 200mmol / L, fully stirred to dissolve, sterilized by filtration, divide...

Embodiment 2

[0046] Example 2: Culture of Breast Cancer Organoids

[0047] 1. Sample acquisition: use an automatic biopsy puncture gun, 18-16G biopsy puncture needle, and the sample length is 15-22 mm. Under the guidance of ultrasound, when reaching the edge of the lesion, start the puncture gun, and quickly pull out the needle after the sample is drawn. For 4-needle samples, the taken tissue is immediately put into the sample preservation solution, stored at 4 degrees, and the container is sealed and transported to the laboratory for further processing within 2 hours.

[0048] 2. Mince the sample to a diameter of 0.5-1 mm in a biological safety cabinet, put it into a centrifuge tube, add 10 ml of the organoid medium A, centrifuge for 10 minutes, and discard the supernatant;

[0049] 3. Add 10 ml of organoid culture medium A and collagenase at a concentration of 2 mg / ml, digest with a shaker at 37 degrees for 2 hours, wash the digested single cells, and count the cells.

[0050] 4. Centri...

Embodiment 3

[0052] Example 3: Passaging of Breast Cancer Organoids

[0053] 1. In a cell culture dish, add 2ml of trypsin to each well, keep it at room temperature for 5 minutes, then pipette with a glass pipette to disperse the cells to form single cells, and transfer them into a centrifuge tube;

[0054] 2. Add 10ml of organoid medium A to the centrifuge tube, centrifuge for 10 minutes, discard the supernatant, count the cells, add 120-240 μl BME, and adjust the cell concentration to 2x10 6 / ml, inoculated in a 24-well plate, 30-60 microliters per well, 200-800 microliters of organoid medium B was added to each well after 20 minutes, and placed in a 37-degree, 5% CO2 cell incubator,

[0055] 3. Change the medium every 3 to 5 days, add 200 to 800 microliters of organoid medium B for each change, and observe it under a microscope. Usually, the passaged breast cancer organoid tissue can be obtained in about 4 weeks. for further research.

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention relates to the fields of cytobiology and cell culture, and particularly relates to a breast cancer organoid culture kit. A method for realizing the kit comprises the following steps: (1)preparing, sub-packaging and using a breast cancer organoid culture medium A; (2) preparing, sub-packaging and using a breast cancer organoid culture medium B; and (3) preparing, sub-packaging and using a sample preserving fluid. The breast cancer organoid culture medium A is a DMEM/F12 culture medium containing 100 U/ml of penicillin, 0.1 mg/ml of streptomycin, 1% of HEPES and 1% of Gluta Max 100 X; the organoid culture medium B is a DMEM/F12 culture medium containing EGF, an FGF growth factor, an ALK inhibitor A83-01, Noggin, an HEPES buffer solution, Y27632 and one or more of a Wnt signalpath related secretory protein family R-Spondin 1-4; and the sample preserving fluid is a DMEM/F12 culture medium containing 100 U/ml of penicillin, 0.1 mg/ml of streptomycin and 1% of L-glutamine. The three components form the breast cancer organoid culture kit.

Description

Technical field: [0001] The invention belongs to the field of cell culture, in particular to a human breast cancer tissue organoid culture kit. Background technique: [0002] Cancer is a major public health problem that seriously threatens people's health. The incidence and mortality of cancer in my country are increasing year by year, causing a major economic burden to families and society, and it is also a major "pain point" for people's livelihood in today's society. Although a large number of anti-tumor drugs are launched every year, the choice of clinical treatment plan mainly depends on various evidence-based guidelines and personal experience of doctors, and cancer is a highly heterogeneous disease. Different patients, different parts, and even internal tumors There is a high degree of diversity signatures that determine cancer progression and response to therapy in different patients. How to better understand the characteristics of tumor heterogeneity, match the bes...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): C12N5/09
CPCC12N5/0693C12N2500/32C12N2501/11C12N2501/115C12N2501/998
Inventor 扈晖郑维越
Owner 扈晖
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap