Department of Pathology and Laboratory Medicine, Weill Cornell Medicine, New York, New York.
Curr Protoc. 2024 Oct;4(10):e70037. doi: 10.1002/cpz1.70037.
Cancer cell lines are important tools to investigate the biology of cancer and test hypotheses to improve cancer treatments. A major challenge in establishing epithelial cancer cell lines is the removal of cancer-associated fibroblasts (CAFs). CAFs are abundant within the tumor microenvironment. CAFs generally proliferate faster than epithelial cancer cells in culture. CAFs can be mistakenly identified as cancer cells, especially when cancer cells display spindle-shaped morphology. Among all cancer types, pancreatic ductal adenocarcinoma (PDAC) is characterized by an abundant desmoplastic stroma. Here, we describe protocols for establishing epithelial cancer cell lines from mouse models of PDAC and verifying that they are not CAFs. The approach is cost-effective and can be used for other types of cancer. If needed, CAF cell lines can also be established and preserved using this protocol. © 2024 Wiley Periodicals LLC. Basic Protocol 1: Isolation of cells from tumors Basic Protocol 2: Isolation and cryopreservation of cancer cell clones Basic Protocol 3: Assessment of the identity of cancer cell lines and CAFs by western blotting.
癌细胞系是研究癌症生物学和检验改善癌症治疗假设的重要工具。建立上皮性癌细胞系的主要挑战之一是去除癌相关成纤维细胞（CAFs）。CAFs 在肿瘤微环境中大量存在。CAFs 在培养中比上皮性癌细胞增殖更快。CAFs 可能会被错误地识别为癌细胞，尤其是当癌细胞呈现出梭形形态时。在所有癌症类型中，胰腺导管腺癌（PDAC）的特征是丰富的纤维母细胞性基质。在这里，我们描述了从小鼠 PDAC 模型中建立上皮性癌细胞系并验证其不是 CAFs 的方案。该方法具有成本效益，可用于其他类型的癌症。如果需要，也可以使用该方案建立和保存 CAF 细胞系。© 2024 Wiley Periodicals LLC. 基本方案 1：从肿瘤中分离细胞 基本方案 2：分离和冷冻保存癌细胞克隆 基本方案 3：通过蛋白质印迹评估癌细胞系和 CAFs 的身份。
