Culture method of breast cancer organoid and co-culture method of tumor-associated fibroblasts

A culture method and breast cancer technology, applied in the field of biomedicine, can solve the problems of expensive culture medium, no fibroblasts, and complicated operation.

Active Publication Date: 2020-11-03
SHANGHAI FIRST PEOPLES HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] However, the medium for the above culture methods is expensive and the operation is cumbersome.
At present, there is no more concise, cheap and efficient breast cancer organoid

Method used

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  • Culture method of breast cancer organoid and co-culture method of tumor-associated fibroblasts
  • Culture method of breast cancer organoid and co-culture method of tumor-associated fibroblasts
  • Culture method of breast cancer organoid and co-culture method of tumor-associated fibroblasts

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Experimental program
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Effect test

Embodiment 1

[0050] 1. Experimental reagents

[0051] Main reagents:

[0052] Reagent name supplier Item No. GlutaMax Gibco 35050061 Collagen type IV Sigma C5139-500MG FBS Gibco 10437028 BME R&D 3433-005-01P Red blood cell lysate Home made Cell recovery solution Corning 354253 TrypLE Express Enzyme Thermo Fisher 12605010 Paraformaldehyde Beyond time P0099-100ml

[0053] PDO complete medium composition and ratio:

[0054]

[0055]

[0056] Washing Buffer formula and ratio: 1x Advanced DMEM / F12, 100xPenicillin / Streptomycin, 0.3% BSA (m / V) (Sigma, B2064), 1000x 20mM EDTA (BioInd, 41-922-25), 1x GlutaMax, 5 μM Y-27632.

[0057] Digesting Buffer formula and ratio: 1x Advanced DMEM / F12, 1mg / mL Collagen type IV, 500μg / mL DNase (Sigma-Aldrich, D5025), 5μM Y-27632, 100xPenicillin / Streptomycin.

[0058] Formula and proportion of freezing solution: 10% DMSO (Sigma-Aldrich, 2650), 90% fetal bovine serum (feta...

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Abstract

The invention relates to a culture method of breast cancer organoids and a co-culture method of tumor-related fibroblasts. According to the invention, a more efficient and convenient improved system for key technical links such as PDO culture, passage, maintenance and the like of tumor tissue organoids of breast cancer patients is established, operation steps and methods for establishing and passage of a PDO cell line are simplified in multiple aspects, components of a culture medium, digestive juice, a washing solution and the like are optimized, and finally, a breast cancer PDO culture system is successfully established. Compared with the prior art, the culture method has the advantages that the success rate and the passage frequency are basically consistent, but the operation is simpler, the required time is shorter, a good foundation is laid for a PDO model to have better clinical and scientific research applications in the future, and the corresponding cost is saved. The inventionalso establishes a stable co-culture system of breast cancer PDO and tumor-associated fibroblasts (CAFs), simultaneous culture, amplification and passage of PDO and CAFs can be realized, and the PDOand the CAFs do not produce obvious influence on each other.

Description

technical field [0001] The invention relates to the field of biomedicine, in particular to a method for culturing breast cancer organoids and a method for co-cultivating tumor-related fibroblasts. Background technique [0002] Breast cancer (Breast Cancer, BC) is the leading cause of death from malignant tumors in women. Globally, China accounts for 12.2% of newly diagnosed cases and 9.6% of BC deaths. According to the latest domestic epidemiological statistics, the incidence of breast cancer ranks first among female malignant tumors, and the mortality rate ranks fifth among female malignant tumors. Breast cancer is a highly heterogeneous cell population that can be divided into several subtypes. Today, breast cancer research relies heavily on established cell lines, which are difficult to maintain in terms of maintaining the cellular biology and behavior of the primary tumor. In clinical treatment, the heterogeneity of breast cancer plays a very important role in guiding ...

Claims

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Application Information

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IPC IPC(8): C12N5/09C12N5/071
CPCC12N5/0693C12N5/0631C12N2502/1323C12N2509/00C12N2509/10C12N2501/11C12N2501/13C12N2501/998C12N2501/15C12N2501/405C12N2500/32C12N2500/38C12N2501/115C12N2513/00Y02A50/30
Inventor 王红霞周天浩鞠高达杜少倩
Owner SHANGHAI FIRST PEOPLES HOSPITAL
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