In the last decade, screening compound libraries on live cells has become an important step in drug discovery. The abundance of compounds in these libraries requires effective high‐throughput (HT) analyzing methods. Although current cell‐based assay protocols are suitable for HT analyses, the analysis itself is often restrained to simple, singular outcomes. Incorporation of HT samplers on flow cytometers has provided an interesting approach to increase the number of measurable parameters and increase the sensitivity and specificity of analyses. Nonetheless, to date, the labor intensive and time‐consuming strategies to detach and stain adherent cells before flow cytometric analysis has restricted use of HT flow cytometry (HTFC) to suspension cells. We have developed a universal “no‐touch” HTFC antibody staining protocol in 384‐well microplates to bypass washing and centrifuging steps of conventional flow cytometry protocols. Optimizing culture conditions, cell‐detachment and staining strategies in 384‐well microplates resulted in an HTFC protocol with an optimal stain index with minimal background staining. The method has been validated using six adherent cell lines and simultaneous staining of four parameters. This HT screening protocol allows for effective monitoring of multiple cellular markers simultaneously, thereby increasing informativity and cost‐effectiveness of drug screening. © 2019 The Authors. Cytometry Part A published by Wiley Periodicals LLC. on behalf of International Society for Advancement of Cytometry.

中文翻译：

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一种贴壁细胞的“非接触”抗体染色方法，用于 384 孔微孔板格式的高通量流式细胞术，用于基于细胞的药物库筛选。

在过去十年中，在活细胞上筛选化合物库已成为药物发现的重要一步。这些文库中的大量化合物需要有效的高通量 (HT) 分析方法。尽管当前基于细胞的检测方案适用于 HT 分析，但分析本身通常仅限于简单、单一的结果。在流式细胞仪上集成 HT 采样器提供了一种有趣的方法来增加可测量参数的数量并提高分析的灵敏度和特异性。尽管如此，迄今为止，在流式细胞术分析之前分离和染色贴壁细胞的劳动密集型和耗时的策略限制了 HT 流式细胞术 (HTFC) 对悬浮细胞的使用。我们在 384 孔微孔板中开发了一种通用的“非接触式”HTFC 抗体染色方案，以绕过传统流式细胞术方案的洗涤和离心步骤。优化 384 孔微孔板中的培养条件、细胞分离和染色策略产生了具有最佳染色指数和最小背景染色的 HTFC 方案。该方法已使用六个贴壁细胞系和四个参数的同时染色进行了验证。这种 HT 筛选方案允许同时有效监测多个细胞标记，从而提高药物筛选的信息量和成本效益。© 2019 作者。384 孔微孔板中的细胞分离和染色策略导致 HTFC 方案具有最佳染色指数和最小背景染色。该方法已使用六个贴壁细胞系和四个参数的同时染色进行了验证。这种 HT 筛选方案允许同时有效监测多个细胞标记，从而提高药物筛选的信息量和成本效益。© 2019 作者。384 孔微孔板中的细胞分离和染色策略导致 HTFC 方案具有最佳染色指数和最小背景染色。该方法已使用六个贴壁细胞系和四个参数的同时染色进行了验证。这种 HT 筛选方案允许同时有效监测多个细胞标记，从而提高药物筛选的信息量和成本效益。© 2019 作者。由 Wiley Periodicals LLC 出版的Cytometry Part A。代表国际细胞计量学促进会。