The rapid advancement of immunotherapy strategies has created a need for technologies that can reliably and reproducibly identify rare populations, detect subtle changes in modulatory signals, and assess antigenic expression patterns that are time-sensitive. Accomplishing these tasks requires careful planning and the employment of tools that provide greater sensitivity and specificity without demanding extensive time. Flow Cytometry has earned its place as a preferred analysis platform. This technology offers a flexible path to the interrogation of protein expression patterns and detection of functional properties in cell populations of interest. Mass Cytometry is a newcomer technology that has generated significant interest in the field. By incorporating mass spectrometry analysis to the traditional principles of flow cytometry, this innovative tool promises to significantly expand the ability to detect multiple proteins on a single cell. The use of these technologies in a manner that is consistent and reproducible through multiple sample sets demands careful attention to experiment design, reagent selection, and instrumentation. Whether applying flow or mass cytometry, reaching successful, reliable results involves many factors. Sample preparation, antibody titrations, and appropriate controls are major biological considerations that impact cytometric analysis. Additionally, instrument voltages, lasers, and run quality assessments are essential for ensuring comparability and reproducibility between analyses. In this article, we aim to discuss the critical aspects that impact flow cytometry, and will touch on important considerations for mass cytometry as well. Focusing on their relevance to immunotherapy studies, we will address the importance of appropriate sample processing and will discuss how selection of suitable panels, controls, and antibodies must follow a carefully designed plan. We will also comment on how educated use of instrumentation plays a significant role in the reliability and reproducibility of results.Through this work, we hope to contribute to the effort toward establishing higher standards for rigor and reproducibility of cytometry practices by researchers, operators, and general cytometry users employing cytometry-based assays in their work. © 2019 International Society for Advancement of Cytometry.

中文翻译：

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免疫肿瘤学细胞计量学实践的严谨性和可重复性：一个多方面的挑战。

免疫治疗策略的快速发展产生了对能够可靠且可重复地识别稀有种群、检测调节信号的细微变化以及评估时间敏感的抗原表达模式的技术的需求。完成这些任务需要仔细规划和使用能够提供更高灵敏度和特异性的工具，而无需花费大量时间。流式细胞术已成为首选分析平台。该技术为询问蛋白质表达模式和检测目标细胞群的功能特性提供了一种灵活的途径。质谱流式细胞术是一项新兴技术，引起了该领域的极大兴趣。通过将质谱分析与流式细胞术的传统原理相结合，这种创新工具有望显着扩大在单个细胞上检测多种蛋白质的能力。以一致且可重复的方式通过多个样品组使用这些技术需要仔细注意实验设计、试剂选择和仪器。无论是应用流式细胞术还是质谱流式细胞术，获得成功、可靠的结果都涉及许多因素。样品制备、抗体滴定和适当的控制是影响细胞计数分析的主要生物学考虑因素。此外，仪器电压、激光和运行质量评估对于确保分析之间的可比性和重现性至关重要。在本文中，我们的目标是讨论影响流式细胞术的关键方面，并将涉及质谱流式细胞术的重要考虑因素。关注它们与免疫治疗研究的相关性，我们将讨论适当样品处理的重要性，并将讨论如何选择合适的面板、对照和抗体必须遵循精心设计的计划。我们还将评论仪器的受过教育的使用如何在结果的可靠性和可重复性方面发挥重要作用。 通过这项工作，我们希望有助于为研究人员、操作员和在他们的工作中使用基于细胞计数的检测的一般细胞计数用户。© 2019 国际细胞计量学促进会。抗体必须遵循精心设计的计划。我们还将评论仪器的受过教育的使用如何在结果的可靠性和可重复性方面发挥重要作用。 通过这项工作，我们希望有助于为研究人员、操作员和在他们的工作中使用基于细胞计数的检测的一般细胞计数用户。© 2019 国际细胞计量学促进会。抗体必须遵循精心设计的计划。我们还将评论仪器的受过教育的使用如何在结果的可靠性和可重复性方面发挥重要作用。 通过这项工作，我们希望有助于为研究人员、操作员和在工作中使用基于细胞术的检测的一般细胞术用户。© 2019 国际细胞计量学促进会。以及在工作中使用基于细胞计数的检测的一般细胞计数用户。© 2019 国际细胞计量学促进会。以及在工作中使用基于细胞计数的检测的一般细胞计数用户。© 2019 国际细胞计量学促进会。