Braakman René B H, Luider Theo M, Martens John W M, Foekens John A, Umar Arzu
Department of Medical Oncology, Center for Translational Molecular Medicine, and Cancer Genomics Centre, Erasmus MC Rotterdam, Rotterdam, The Netherlands.
Methods Mol Biol. 2011;755:143-54. doi: 10.1007/978-1-61779-163-5_11.
Breast cancer tissues are characterized by cellular heterogeneity, representing a mixture of, e.g., healthy epithelial ducts, invasive or in situ tumor cells, surrounding stroma, infiltrating immune cells, blood vessels, and capillaries. As a consequence, protein extracts from whole tissue lysates also represent a variety of cell types present in the tissues under examination. This, however, seriously hampers the analysis of tumor cell-specific signals, which is of interest when performing biomarker discovery-type of studies. Therefore, laser capture microdissection is a perfect tool to isolate a relatively pure population of cells of interest, such as tumor cells. In this chapter, we describe the use of the PALM MicroBeam system for laser microdissection and pressure catapulting. Protocols are provided for sectioning, staining, microdissection, sample preparation, and mass spectrometric analysis of snap frozen breast cancer tissue.
乳腺癌组织具有细胞异质性，例如，它是由健康上皮导管、浸润性或原位肿瘤细胞、周围基质、浸润免疫细胞、血管和毛细血管等混合而成。因此，来自全组织裂解物的蛋白质提取物也代表了所检测组织中存在的多种细胞类型。然而，这严重阻碍了肿瘤细胞特异性信号的分析，而在进行生物标志物发现类研究时，这种分析是很有意义的。因此，激光捕获显微切割是分离相对纯净的目标细胞群体（如肿瘤细胞）的理想工具。在本章中，我们描述了使用PALM微束系统进行激光显微切割和压力弹射的方法。文中提供了关于速冻乳腺癌组织的切片、染色、显微切割、样品制备和质谱分析的实验方案。
