We describe a sensitive, robust, high-throughput method for quantifying the ability of metastatic tumor cells to colonize a secondary organ. Metastasis is the leading cause of death in cancer patients, and successful colonization of the secondary organ is the rate-limiting step in the metastatic process; thus, experimental methods that can be used to interrogate the key factors required for this critical step are of great importance. The experimental metastasis assay we detail here includes tail-vein injection of cancer cells into the mouse and determination of the resulting secondary organ colonization, primarily in the lung, 10 d post dosing. This assay can be used to investigate factors that regulate metastatic colonization both at the tumor-cell-intrinsic level (via manipulation of the tumor cells before injection) and at the tumor-cell-extrinsic level (such as the tissue microenvironment, via the use of genetically modified (GM) mice or agents such as antibodies or drugs). Using this method, we have robustly screened more than 950 GM mouse lines to identify novel microenvironmental regulators of metastatic colonization. The experimental details discussed here include choosing of appropriate cell numbers, handling of the cells, selection of recipient animals and injection techniques. Furthermore, we discuss key experimental design considerations, including the choice of the method used to determine metastatic burden and statistical analysis of the results, as well as provide troubleshooting tips and identification of factors that contribute to experimental variability.

中文翻译：

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一种用于鉴定转移性定植调节剂的小鼠体内高通量筛选方法。

我们描述了一种灵敏、稳健、高通量的方法，用于量化转移性肿瘤细胞定植次级器官的能力。转移是癌症患者死亡的主要原因，第二器官的成功定植是转移过程中的限速步骤；因此，可用于询问这一关键步骤所需关键因素的实验方法非常重要。我们在这里详细介绍的实验性转移测定包括将癌细胞尾静脉注射到小鼠体内，并在给药后 10 天确定由此产生的次级器官定植，主要是在肺中。该测定可用于研究在肿瘤细胞内在水平（通过在注射前对肿瘤细胞进行操作）和在肿瘤细胞外在水平（例如组织微环境，通过使用转基因 (GM) 小鼠或抗体或药物等试剂）。使用这种方法，我们稳健地筛选了 950 多个 GM 小鼠系，以识别转移性定植的新型微环境调节剂。这里讨论的实验细节包括选择合适的细胞数量、细胞的处理、受体动物的选择和注射技术。此外，我们讨论了关键的实验设计考虑因素，包括用于确定转移负担的方法的选择和结果的统计分析，